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Bellows for consant-velocity joints

A convoluted boot for sealing an annular gap between two parts which are connected to one another in a rotationally fast way, which can be articulated relative to one another and which are axially displaceable relative to one another, especially of a constant velocity plunging joint, consisting of a low-strain polymer hard material, having a first larger collar 11 to be secured to a first component, a second smaller collar 12 to be secured to a second component and a plurality of annular fold units which extend between the first collar and the second collar and which, in the form of outer folds consisting of two annular flanks 31, 32, form a fold peak 21 between two fold valleys 22, in a first group A of at least three annular folds with a connection to the first collar 11, the diameters of the fold peaks 21 and fold valleys 22 decrease in the direction from the first collar to the second collar, in a second group B of annular folds with a connection to the second collar 12 of at least one fold, the diameters of the fold valleys and optionally of the fold peaks 21 are constant, the ratio of the diameter D1 of the fold peak 21 of the largest annular fold A1 of the first group A to the diameter D2 of the second collar 12 amounts to ≧2.5.

1. A convoluted boot for sealing an annular gap between two parts which are connected to one another in a rotationally fast way, which can be articulated relative to one another and which are axially displaceable relative to one another, the boot comprising: a low-strain hard polymer material, having a first larger collar (11) to be secured to a first component, a second smaller collar (12) to be secured to a second component and a plurality of annular fold units which extend between the first collar and the second collar and which, in the form of outer folds comprising two annular flanks (31, 32), form a fold peak (21) between two fold valleys (22); wherein, in a first group (A) of at least three annular folds with a connection to the first collar (11), the diameters of the fold peaks (21) and fold valleys (22) decrease from the first collar to the second collar, and wherein in a second group (B) of at least one annular fold with a connection to the second collar (12), the diameters of the fold valleys are constant; and wherein the ratio of the diameter (D1) of the fold peak (21) of the largest annular fold (A1) of the first group (A) to the diameter (D2) of the second collar (12) is greater than or equal to 2.5 and the two annular flanks (31, 32) of each of the annular folds of the first group (A) form opposed angles with a radial plane (R), wherein a smaller angle (β) is formed by the annular flank (32) pointing towards the second collar (12) and wherein a larger angle (α) is formed by the annular flank pointing towards the first collar (11), the larger angle (α) being at least 25° greater than the smaller angle (β). 2. A convoluted boot according to claim 1, wherein the first group (A) comprises up to five annular folds. 3. A convoluted boot according to claim 1 wherein the second group (B) comprises up to eight annular folds. 4. A convoluted boot according to claim 1 wherein the annular flanks (31, 32) of each of the annular folds of the second group (B) form opposed angles with a radial plane, wherein the annular flank (32) pointing towards the second collar (12) forms first angle (β) and wherein the annular flank (31) pointing towards first collar (11) forms a second angle (α) being in a range from 5° smaller to 5° greater than the first angle (β). 5. A convoluted boot according to claim 1 wherein, between the annular folds of the first group (A) and the annular folds of the second group (B), there are provided transition folds (C) whose diameters deviate from one another at the fold valleys (22), wherein the diameter of the fold valley at the annular flank (31) pointing towards the first collar (11) is greater than the diameter of the fold valley at the annular flank (32) pointing towards the second collar (12). 6. A convoluted boot according to claim 5, wherein the annular flanks (31, 32) of the transition fold (C) form opposed angles with a radial plane (R), wherein the annular flank (32) pointing towards the second collar (12) forms a first angle (β) and that the annular flank (31) pointing towards the first collar (11) forms a second angle (α) which is defined by (β+25°)≧α≧(β+5°). 7. A convoluted boot according to claim 1 wherein the convoluted boot comprises a thermoplastic elastomer (TPE). 8. A convoluted boot according to claim 7, wherein the TPE comprises a polyurethane (TPU). 9. A convoluted boot according to claim 7, wherein the TPE comprises a polyester (TPEE). 10. A convoluted boot according to claim 9, wherein the TPEE is a polyether ester or a polyester ester. 11. A convoluted boot according to claim 7, wherein the TPE comprises a polyamide (TPA). 12. A convoluted boot according to claim 7, wherein the TPE comprises a polyolefin (TPO). 13. A convoluted boot according to claim 12, wherein the TPO is a polypropylene or a polyethylene. 14. A convoluted boot according to claim 1 wherein the diameters of the fold peaks (21) of the second group (B) of annular folds are constant. 15. A convoluted boot according to claim 2 wherein the second group (B) comprises up to eight annular folds. 16. A convoluted boot according to claim 2 wherein the annular flanks (31, 32) of each of the annular folds of the second group (B) form opposed angles with a radial plane, wherein the annular flank (32) pointing towards the second collar (12) forms first angle (β) and wherein the annular flank (31) pointing towards first collar (11) forms a second angle (α) which differs from the first angle (β) not more than 5°. 17. A convoluted boot according to claim 3 wherein the annular flanks (31, 32) of each of the annular folds of the second group (B) form opposed angles with a radial plane, wherein the annular flank (32) pointing towards the second collar (12) forms first angle (β) and wherein the annular flank (31) pointing towards first collar (11) forms a second angle (α) which differs from the first angle (β) not more than 5°. 18. A convoluted boot according to claim 2 wherein, between the annular folds of the first group (A) and the annular folds of the second group (B), there is provided at least one transition fold (C) whose diameters deviate from one another at the fold valleys (22), wherein the diameter of the fold valley at the annular flank (31) pointing towards the first collar (11) is greater than the diameter of the fold valley at the annular flank (32) pointing towards the second collar (12). 19. A convoluted boot according to claim 3 wherein, between the annular folds of the first group (A) and the annular folds of the second group (B), there is provided at least one transition fold (C) whose diameters deviate from one another at the fold valleys (22), wherein the diameter of the fold valley at the annular flank (31) pointing towards the first collar (11) is greater than the diameter of the fold valley at the annular flank (32) pointing towards the second collar (12). 20. A convoluted boot according to claim 4 wherein, between the annular folds of the first group (A) and the annular folds of the second group (B), there is provided at least one transition fold (C) whose diameters deviate from one another at the fold valleys (22), wherein the diameter of the fold valley at the annular flank (31) pointing towards the first collar (11) is greater than the diameter of the fold valley at the annular flank (32) pointing towards the second collar (12). 21. A convoluted boot according to claim 16 wherein, between the annular folds of the first group (A) and the annular folds of the second group (B), there is provided at least one transition fold (C) whose diameters deviate from one another at the fold valleys (22), wherein the diameter of the fold valley at the annular flank (31) pointing towards the first collar (11) is greater than the diameter of the fold valley at the annular flank (32) pointing towards the second collar (12). 22. A convoluted boot according to claim 7 wherein, between the annular folds of the first group (A) and the annular folds of the second group (B), there is provided at least one transition fold (C) whose diameters deviate from one another at the fold valleys (22), wherein the diameter of the fold valley at the annular flank (31) pointing towards the first collar (11) is greater than the diameter of the fold valley at the annular flank (32) pointing towards the second collar (12). 23. A convoluted boot according to claim 18, wherein the annular flanks (31, 32) of the transition fold (C) form opposed angles with a radial plane (R), wherein the annular flank (32) pointing towards the second collar (12) forms a first angle (β) and that the annular flank (31) pointing towards the first collar (11) forms a second angle (α) which is defined by (β+25°)≧α≧(β+5°). 24. A convoluted boot according to claim 19, wherein the annular flanks (31, 32) of the transition fold (C) form opposed angles with a radial plane (R), wherein the annular flank (32) pointing towards the second collar (12) forms a first angle (β) and that the annular flank (31) pointing towards the first collar (11) forms a second angle (α) which is defined by (β+25°)≧α≧(β+5°). 25. A convoluted boot according to claim 20, wherein the annular flanks (31, 32) of the transition fold (C) form opposed angles with a radial plane (R), wherein the annular flank (32) pointing towards the second collar (12) forms a first angle (β) and that the annular flank (31) pointing towards the first collar (11) forms a second angle (α) which is defined by (β+25°)≧α≧(β+5°). 26. A convoluted boot according to claim 21, wherein the annular flanks (31, 32) of the transition fold (C) form opposed angles with a radial plane (R), wherein the annular flank (32) pointing towards the second collar (12) forms a first angle (β) and that the annular flank (31) pointing towards the first collar (11) forms a second angle (α) which is defined by (β+25°)≧α≧(β+5°). 27. A convoluted boot according to claim 22, wherein the annular flanks (31, 32) of the transition fold (C) form opposed angles with a radial plane (R), wherein the annular flank (32) pointing towards the second collar (12) forms a first angle (β) and that the annular flank (31) pointing towards the first collar (11) forms a second angle (α) which is defined by (β+25°)≧α≧(β+5°). 28. A convoluted boot according to claim 27 wherein the convoluted boot comprises a thermoplastic elastomer (TPE).

Novel nucleic acids and polypeptides

The present invention provides novel nucleic acids, novel polypeptide sequences encoded by these nucleic acids and uses thereof.

1. An isolated polynucleotide comprising a nucleotide sequence selected from the group consisting of SEQ ID NO: 1-8051, a mature protein coding portion of SEQ ID NO: 1-8051, an active domain of SEQ ID NO: 1-8051, and complementary sequences thereof. 2. An isolated polynucleotide encoding a polypeptide with biological activity, wherein said polynucleotide hybridizes to the polynucleotide of claim 1 under stringent hybridization conditions. 3. An isolated polynucleotide encoding a polypeptide with biological activity, wherein said polynucleotide has greater than about 90% sequence identity with the polynucleotide of claim 1. 4. The polynucleotide of claim 1 wherein said polynucleotide is DNA. 5. An isolated polynucleotide of claim 1 wherein said polynucleotide comprises the complementary sequences. 6. A vector comprising the polynucleotide of claim 1. 7. An expression vector comprising the polynucleotide of claim 1. 8. A host cell genetically engineered to comprise the polynucleotide of claim 1. 9. A host cell genetically engineered to comprise the polynucleotide of claim 1 operatively associated with a regulatory sequence that modulates expression of the polynucleotide in the host cell. 10. An isolated polypeptide, wherein the polypeptide is selected from the group consisting of: (a) a polypeptide encoded by any one of the polynucleotides of claim 1; and (b) a polypeptide encoded by a polynucleotide hybridizing under stringent conditions with any one of SEQ ID NO: 1-8051. 11. A composition comprising the polypeptide of claim 10 and a carrier. 12. An antibody directed against the polypeptide of claim 10. 13. A method for detecting the polynucleotide of claim 1 in a sample, comprising: a) contacting the sample with a compound that binds to and forms a complex with the polynucleotide of claim 1 for a period sufficient to form the complex; and b) detecting the complex, so that if a complex is detected, the polynucleotide of claim 1 is detected. 14. A method for detecting the polynucleotide of claim 1 in a sample, comprising: a) contacting the sample under stringent hybridization conditions with nucleic acid primers that anneal to the polynucleotide of claim 1 under such conditions; b) amplifying a product comprising at least a portion of the polynucleotide of claim 1; and c) detecting said product and thereby the polynucleotide of claim 1 in the sample. 15. The method of claim 14, wherein the polynucleotide is an RNA molecule and the method further comprises reverse transcribing an annealed RNA molecule into a cDNA polynucleotide. 16. A method for detecting the polypeptide of claim 10 in a sample, comprising: a) contacting the sample with a compound that binds to and forms a complex with the polypeptide under conditions and for a period sufficient to form the complex; and b) detecting formation of the complex, so that if a complex formation is detected, the polypeptide of claim 10 is detected. 17. A method for identifying a compound that binds to the polypeptide of claim 10, comprising: a) contacting the compound with the polypeptide of claim 10 under conditions sufficient to form a polypeptide/compound complex; and b) detecting the complex, so that if the polypeptide/compound complex is detected, a compound that binds to the polypeptide of claim 10 is identified. 18. A method for identifying a compound that binds to the polypeptide of claim 10, comprising: a) contacting the compound with the polypeptide of claim 10, in a cell, under conditions sufficient to form a polypeptide/compound complex, wherein the complex drives expression of a reporter gene sequence in the cell; and b) detecting the complex by detecting reporter gene sequence expression, so that if the polypeptide/compound complex is detected, a compound that binds to the polypeptide of claim 10 is identified. 19. A method of producing the polypeptide of claim 10, comprising, a) culturing a host cell comprising a polynucleotide sequence selected from the group consisting of a polynucleotide sequence of SEQ ID NO: 1-8051, a mature protein coding portion of SEQ ID NO: 1-8051, an active domain of SEQ ID NO: 1-8051, complementary sequences thereof and a polynucleotide sequence hybridizing under stringent conditions to SEQ ID NO: 1-8051, under conditions sufficient to express the polypeptide in said cell; and b) isolating the polypeptide from the cell culture or cells of step (a). 20. An isolated polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 8052-16102, the mature protein portion thereof, or the active domain thereof. 21. The polypeptide of claim 20 wherein the polypeptide is provided on a polypeptide array. 22. A collection of polynucleotides, wherein the collection comprises the sequence information of at least one of SEQ ID NO: 1-8051. 23. The collection of claim 22, wherein the collection is provided on a nucleic acid array. 24. The collection of claim 23, wherein the array detects full-matches to any one of the polynucleotides in the collection. 25. The collection of claim 23, wherein the array detects mismatches to any one of the polynucleotides in the collection. 26. The collection of claim 22, wherein the collection is provided in a computer-readable format. 27. A method of treatment comprising administering to a mammalian subject in need thereof a therapeutic amount of a composition comprising a polypeptide of claim 10 or 20 and a pharmaceutically acceptable carrier. 28. A method of treatment comprising administering to a mammalian subject in need thereof a therapeutic amount of a composition comprising an antibody that specifically binds to a polypeptide of claim 10 or 20 and a pharmaceutically acceptable carrier.

<SOH> 2. BACKGROUND <EOH>Technology aimed at the discovery of protein factors (including e.g., cytokines, such as lymphokines, interferons, circulating soluble factors, chemokines, and interleukins) has matured rapidly over the past decade. The now routine hybridization cloning and expression cloning techniques clone novel polynucleotides “directly” in the sense that they rely on information directly related to the discovered protein (i.e., partial DNA/amino acid sequence of the protein in the case of hybridization cloning; activity of the protein in the case of expression cloning). More recent “indirect” cloning techniques such as signal sequence cloning, which isolates DNA sequences based on the presence of a now well-recognized secretory leader sequence motif, as well as various PCR-based or low stringency hybridization-based cloning techniques, have advanced the state of the art by making available large numbers of DNA/amino acid sequences for proteins that are known to have biological activity, for example, by virtue of their secreted nature in the case of leader sequence cloning, by virtue of their cell or tissue source in the case of PCR-based techniques, or by virtue of structural similarity to other genes of known biological activity. Identified polynucleotide and polypeptide sequences have numerous applications in, for example, diagnostics, forensics, gene mapping; identification of mutations responsible for genetic disorders or other traits, to assess biodiversity, and to produce many other types of data and products dependent on DNA and amino acid sequences.

<SOH> 3. SUMMARY OF THE INVENTION <EOH>The compositions of the present invention include novel isolated polypeptides, novel isolated polynucleotides encoding such polypeptides, including recombinant DNA molecules, cloned genes or degenerate variants thereof, especially naturally occurring variants such as allelic variants, antisense polynucleotide molecules, and antibodies that specifically recognize one or more epitopes present on such polypeptides, as well as hybridomas producing such antibodies. The compositions of the present invention additionally include vectors, including expression vectors, containing the polynucleotides of the invention, cells genetically engineered to contain such polynucleotides and cells genetically engineered to express such polynucleotides. The present invention relates to a collection or library of at least one novel nucleic acid sequence assembled from expressed sequence tags (ESTs) isolated mainly by sequencing by hybridization (SBH), and in some cases, sequences obtained from one or more public databases. The invention relates also to the proteins encoded by such polynucleotides, along with therapeutic, diagnostic and research utilities for these polynucleotides and proteins. These nucleic acid sequences are designated as SEQ ID NO: 1-8051. The polypeptides sequences are designated SEQ ID NO: 8052-16102. The nucleic acids and polypeptides are provided in the Sequence Listing. In the nucleic acids provided in the Sequence Listing, A is adenosine; C is cytosine; G is guanine; T is thymine; and N is any of the four bases. In the amino acids provided in the Sequence Listing, * corresponds to the stop codon. The nucleic acid sequences of the present invention also include, nucleic acid sequences that hybridize to the complement of SEQ ID NO: 1-8051 under stringent hybridization conditions; nucleic acid sequences which are allelic variants or species homologues of any of the nucleic acid sequences recited above, or nucleic acid sequences that encode a peptide comprising a specific domain or truncation of the peptides encoded by SEQ ID NO: 1-8051. A polynucleotide comprising a nucleotide sequence having at least 90% identity to an identifying sequence of SEQ ID NO: 1-8051 or a degenerate variant or fragment thereof. The identifying sequence can be 100 base pairs in length. The nucleic acid sequences of the present invention also include the sequence information from the nucleic acid sequences of SEQ ID NO: 1-8051. The sequence information can be a segment of any one of SEQ ID NO: 1-8051 that uniquely identifies or represents the sequence information of SEQ ID NO: 1-8051. A collection as used in this application can be a collection of only one polynucleotide. The collection of sequence information or identifying information of each sequence can be provided on a nucleic acid array. In one embodiment, segments of sequence information is provided on a nucleic acid array to detect the polynucleotide that contains the segment. The array can be designed to detect full-match or mismatch to the polynucleotide that contains the segment. The collection can also be provided in a computer-readable format. This invention also includes the reverse or direct complement of any of the nucleic acid sequences recited above; cloning or expression vectors containing the nucleic acid sequences; and host cells or organisms transformed with these expression vectors. Nucleic acid sequences (or their reverse or direct complements) according to the invention have numerous applications in a variety of techniques known to those skilled in the art of molecular biology, such as use as hybridization probes, use as primers for PCR, use in an array, use in computer-readable media, use in sequencing full-length genes, use for chromosome and gene mapping, use in the recombinant production of protein, and use in the generation of anti-sense DNA or RNA, their chemical analogs and the like. In a preferred embodiment, the nucleic acid sequences of SEQ ID NO: 1-8051 or novel segments or parts of the nucleic acids of the invention are used as primers in expression assays that are well known in the art. In a particularly preferred embodiment, the nucleic acid sequences of SEQ ID NO: 1-8051 or novel segments or parts of the nucleic acids provided herein are used in diagnostics for identifying expressed genes or, as well known in the art and exemplified by Vollrath et al., Science 258:52-59 (1992), as expressed sequence tags for physical mapping of the human genome. The isolated polynucleotides of the invention include, but are not limited to, a polynucleotide comprising any one of the nucleotide sequences set forth in SEQ ID NO: 1-8051; a polynucleotide comprising any of the full length protein coding sequences of SEQ ID NO: 1-8051; and a polynucleotide comprising any of the nucleotide sequences of the mature protein coding sequences of SEQ ID NO: 1-8051. The polynucleotides of the present invention also include, but are not limited to, a polynucleotide that hybridizes under stringent hybridization conditions to (a) the complement of any one of the nucleotide sequences set forth in SEQ ID NO: 1-8051; (b) a nucleotide sequence encoding any one of the amino acid sequences set forth in the Sequence Listing (e.g., SEQ ID NO: 8052-16102); (c) a polynucleotide which is an allelic variant of any polynucleotides recited above; (d) a polynucleotide which encodes a species homolog (e.g. orthologs) of any of the proteins recited above; or (e) a polynucleotide that encodes a polypeptide comprising a specific domain or truncation of any of the polypeptides comprising an amino acid sequence set forth in the Sequence Listing. The isolated polypeptides of the invention include, but are not limited to, a polypeptide comprising any of the amino acid sequences set forth in the Sequence Listing; or the corresponding full length or mature protein. Polypeptides of the invention also include polypeptides with biological activity that are encoded by (a) any of the polynucleotides having a nucleotide sequence set forth in SEQ ID NO: 1-8051; or (b) polynucleotides that hybridize to the complement of the polynucleotides of (a) under stringent hybridization conditions. Biologically or immunologically active variants of any of the polypeptide sequences in the Sequence Listing, and “substantial equivalents” thereof (e.g., with at least about 65%, 70%, 75%, 80%, 85%, 90%, 95%, 98% or 99% amino acid sequence identity) that preferably retain biological activity are also contemplated. The polypeptides of the invention may be wholly or partially chemically synthesized but are preferably produced by recombinant means using the genetically engineered cells (e.g. host cells) of the invention. The invention also provides compositions comprising a polypeptide of the invention. Polypeptide compositions of the invention may further comprise an acceptable carrier, such as a hydrophilic, e.g., pharmaceutically acceptable, carrier. The invention also provides host cells transformed or transfected with a polynucleotide of the invention. The invention also relates to methods for producing a polypeptide of the invention comprising growing a culture of the host cells of the invention in a suitable culture medium under conditions permitting expression of the desired polypeptide, and purifying the polypeptide from the culture or from the host cells. Preferred embodiments include those in which the protein produced by such process is a mature form of the protein. Polynucleotides according to the invention have numerous applications in a variety of techniques known to those skilled in the art of molecular biology. These techniques include use as hybridization probes, use as oligomers, or primers, for PCR, use for chromosome and gene mapping, use in the recombinant production of protein, and use in generation of anti-sense DNA or RNA, their chemical analogs and the like. For example, when the expression of an mRNA is largely restricted to a particular cell or tissue type, polynucleotides of the invention can be used as hybridization probes to detect the presence of the particular cell or tissue mRNA in a sample using, e.g., in situ hybridization. In other exemplary embodiments, the polynucleotides are used in diagnostics as expressed sequence tags for identifying expressed genes or, as well known in the art and exemplified by Vollrath et al., Science 258:52-59 (1992), as expressed sequence tags for physical mapping of the human genome. The polypeptides according to the invention can be used in a variety of conventional procedures and methods that are currently applied to other proteins. For example, a polypeptide of the invention can be used to generate an antibody that specifically binds the polypeptide. Such antibodies, particularly monoclonal antibodies, are useful for detecting or quantitating the polypeptide in tissue. The polypeptides of the invention can also be used as molecular weight markers, and as a food supplement. Methods are also provided for preventing, treating, or ameliorating a medical condition which comprises the step of administering to a mammalian subject a therapeutically effective amount of a composition comprising a polypeptide of the present invention and a pharmaceutically acceptable carrier. In particular, the polypeptides and polynucleotides of the invention can be utilized, for example, in methods for the prevention and/or treatment of disorders involving aberrant protein expression or biological activity. The present invention further relates to methods for detecting the presence of the polynucleotides or polypeptides of the invention in a sample. Such methods can, for example, be utilized as part of prognostic and diagnostic evaluation of disorders as recited herein and for the identification of subjects exhibiting a predisposition to such conditions. The invention provides a method for detecting the polynucleotides of the invention in a sample, comprising contacting the sample with a compound that binds to and forms a complex with the polynucleotide of interest for a period sufficient to form the complex and under conditions sufficient to form a complex and detecting the complex such that if a complex is detected, the polynucleotide of interest is detected. The invention also provides a method for detecting the polypeptides of the invention in a sample comprising contacting the sample with a compound that binds to and forms a complex with the polypeptide under conditions and for a period sufficient to form the complex and detecting the formation of the complex such that if a complex is formed, the polypeptide is detected. The invention also provides kits comprising polynucleotide probes and/or monoclonal antibodies, and optionally quantitative standards, for carrying out methods of the invention. Furthermore, the invention provides methods for evaluating the efficacy of drugs, and monitoring the progress of patients, involved in clinical trials for the treatment of disorders as recited above. The invention also provides methods for the identification of compounds that modulate (i.e., increase or decrease) the expression or activity of the polynucleotides and/or polypeptides of the invention. Such methods can be utilized, for example, for the identification of compounds that can ameliorate symptoms of disorders as recited herein. Such methods can include, but are not limited to, assays for identifying compounds and other substances that interact with (e.g., bind to) the polypeptides of the invention. The invention provides a method for identifying a compound that binds to the polypeptides of the invention comprising contacting the compound with a polypeptide of the invention in a cell for a time sufficient to form a polypeptide/compound complex, wherein the complex drives expression of a reporter gene sequence in the cell; and detecting the complex by detecting the reporter gene sequence expression such that if expression of the reporter gene is detected the compound that binds to a polypeptide of the invention is identified. The methods of the invention also provides methods for treatment which involve the administration of the polynucleotides or polypeptides of the invention to individuals exhibiting symptoms or tendencies. In addition, the invention encompasses methods for treating diseases or disorders as recited herein comprising administering compounds and other substances that modulate the overall activity of the target gene products. Compounds and other substances can effect such modulation either on the level of target gene/protein expression or target protein activity. The polypeptides of the present invention and the polynucleotides encoding them are also useful for the same functions known to one of skill in the art as the polypeptides and polynucleotides to which they have homology (set forth in the sequence listing). If no homology is set forth for a sequence, then the polypeptides and polynucleotides of the present invention are useful for a variety of applications, as described herein, including use in arrays for detection. detailed-description description="Detailed Description" end="lead"?

Combinations of enzyme inhibitor-containing preparations and the use thereof

The invention comprises a process wherein the DNA synthesis and, thus, the proliferation of mononuclear cells (MNZ) and of T cells as well is inhibited by the simultaneous and combined inhibition of the enzyme activity (I) of alanyl aminopeptidase and of dipeptidyl peptidase IV; (II) of dipeptidyl peptidase IV and of the angiotensin-converting enzyme; (III) of dipeptidyl peptidase IV and of prolyl oligopeptidase; and (IV) of dipeptidyl peptidase IV and of X-Pro-aminopeptidase to an extent which cannot be achieved by an application of a single one of said enzyme inhibitors, even at a higher dosage. Although the said inhibitors exercise an influence on the very same process finally, i.e. the DNA synthesis and, thus, the proliferation of immune cells, this effect is not complete and is not long lasting when a single inhibitor is applied. From the functional overlap of enzymatic activities results an additive/superadditive inhibitory effect on the DNA synthesis and proliferation by the simultaneous inhibition of more than one of the above enzymes, as our data show. Our invention shows that the simultaneous application of substances inhibiting the above enzymes or of corresponding preparations and administration forms, respectively, is well suitable for a therapy of autoimmune diseases and chronic diseases with an inflammatory genesis as well as for a treatment of rejection episodes after a transplantation.

1. Use of inhibitors of dipeptidyl peptidase IV (DP IV) as well as of enzymes having the same substrate specificity (DP IV-analogous enzyme activity) in combination with inhibitors of alanyl aminopeptidase (aminopeptidase N, APN) as well as of enzymes having the same substrate specificity (APN-analogous enzyme activity), of X-Pro-aminopeptidase (aminopeptidase P, APP), of the angiotensin-converting enzyme (ACE) and/or of prolyl oligopeptidase (POP, prolyl endopeptidase, PEP) for a superadditive inhibition of an activation, DNA synthesis and proliferation of human T lymphocytes and mononuclear cells. 2. Use according to claim 1, wherein the inhibitors of DP IV are preferably Xaa-Pro-dipeptides (Xaa=α-amino acid or side chain-protected derivative), corresponding derivatives, preferably dipeptide phosphonic acid diaryl estates, dipeptide boronic acids (e.g. Pro-boro-Pro) and salts thereof, Xaa-Xaa-(Trp)-Pro-(Xaa)n-peptides (Xaa=α-amino acid, n=0-10), corresponding derivatives and salts thereof or amino acid (Xaa)-amides, corresponding derivatives and salts thereof, wherein Xaa is an α-amino acid or a side chain-protected derivative, respectively, preferably NE-4-nitrobenzyloxycarbonyl L-lysine, -L-proline, -L-tryptophane, -L-isoleucine, -L-valine and cyclic amines as, for example, pyrrolidine, piperidine, thiazolidine and derivatives thereof act as amide structure. 3. Use according to claim 1, wherein amino acid amides, e.g., NE-4-nitrobenzyloxy-carbonyl-L-lysine-thiazolidid, -pyrrolidid and -piperidid as well as the corresponding 2-cyanothiazolidid, 2-cyanopyrrolidid and 2-cyanopiperidid derivative, are preferably employed as the DP IV inhibitor. 4. Use according to claim 1, wherein Actinonin, Leuhistin, Phebestin, Amastin, Probestin, β-aminothiols, α-amino phosphinic acids, α-amino phosphinic acid derivatives, preferably D-Phe-ψ-[PO(OH)—CH2]-Phe-Phe and salts thereof, preferably act as inhibitors of APN; Apstatin, (2S,3R)-HAMH-L-proline, (2S,3R)-HAPB-L-proline, the corresponding L-proline methyl esters, (2S,3R)-HAMH/(2S,3R)-HAPB-pyrrolidides, -thiazolidides (HAMH=3-amino-2-hydroxy-5-methyl hexanoyl, HAPB=3-amino-2-hydroxy-4-phenyl butanoyl) and salts thereof preferably act as inhibitors of APP; Captopril, Enalapril, Lisinopril, Cilazopril and salts thereof preferably act as inhibitors of ACE; Postatin, Eurystatin A or B, Na-protected peptide aldehydes, preferably benzyloxy-carbonyl-L-prolyl-L-prolinal or benzyloxycarbonyl-L-thioprolyl-L-thioprolinal, Na-protected amino acid (Xaa)-pyrrolidides or -thiazolidides (Xaa=α-amino acid, preferably L-alanine, L-valine, L-isoleucine) as well as the corresponding 2-cyano-pyrrolidid or 2-cyanothiazolidid derivatives, substrate-analogous Na-protected peptide phosphonic acid diaryl esters or peptide diazo methyl ketones or peptide ammonium methyl ketones and salts thereof preferably act as inhibitors of POP (PEP). 5. Use of inhibitor combinations of claim 1 for a prevention and therapy of autoimmune diseases, preferably of Rheumatoid Arthritis, Lupus Erythematodes, Multiple Sclerosis, IDDM, Morbus Crohn, Colitis Ulcerosa, Psoriasis, Neurodermitis, Glomerulonephritis, Interstitial Nephritis, Vasculitis, autoimmune diseases of the thyroid gland or autoimmune hemolytic anemia as well as of other chronic diseases with an inflammatory genesis as, for example, allergies and arteriosclerosis. 6. Use of inhibitor combinations of claim 1 for a suppression of transplant rejection and for a therapy of tumor diseases. 7. Pharmaceutical preparations, comprising inhibitors of dipeptidyl peptidase IV (DP IV) as well as of enzymes having DP IV-analogous enzyme activity in combination with inhibitors of any of the enzymes alanyl aminopeptidase (aminopeptidase N, APN) as well as of enzymes having the same substrate specificity (APN-analogous enzyme activity), of X-Pro aminopeptidase (aminopeptidase P, APP), of the angiotensin-converting enzyme (ACE) and of prolyl oligopeptidase (POP, prolyl endopeptidase, PEP) and in combination with per se known carrier, additive and/or auxiliary substances. 8. Pharmaceutical preparation according to claim 7, comprising preferably Xaa-Pro-dipeptides (Xaa=α-amino acid or side chain-protected derivatives) corresponding derivatives, preferably dipeptide phosphonic acid diaryl esters and salts thereof, Xaa-Xaa-(Trp)-Pro-(Xaa)n-peptides (Xaa=α-amino acid, n=0-10), corresponding derivatives and salts thereof or amino acid (Xaa)-amides, corresponding derivatives and salts thereof, wherein Xaa is an α-amino acid or a side chain-protected derivative, respectively, preferably Ne-4-nitrobenzyloxcarbonyl L-lysine, -L-proline, -L-tryptophane, -L-isoleucine, -L-valine, and cyclic amines as, for example, pyrrolidine, piperidine, thiazolidine and derivatives thereof act as amide structure, as the inhibitors of DP IV. 9. Pharmaceutical preparation according to claim 7, comprising preferably amino acid amides, e.g. Ne-4-nitrobenzyloxcarbonyl-L-lysine-thiazolidid, -pyrrolidid and -piperidid as well as the corresponding 2-cyanothiazolidid, 2-cyanopyrrolidid and 2-cyanopiperidid derivative as the inhibitors of DP IV. 10. Pharmaceutical preparation according to claim 7, comprising, as the inhibitors of APN, APP, ACE and POP (PEP), preferably Actnonin, Leuhistin, Phebestin, Amastin, Probestin, β-aminothiols, α-amino phosphinic acids, α-amino phoshinic acid derivatives, preferably D-Phe-ψ-[PO(OH)—CH2]-Phe-Phe and salts thereof, as the inhibitors of APN; Apstatin, (2S,3R)-HAMH-L-proline, (2S,3R)-HAPB-pyrrolidides, -thiazolidides (HAMH=3-amino-2-hydroxy-5-methyl hexanoyl, HAPB=3-amino-2-hydroxy-4-phenyl butanoyl) and salts thereof as the inhibitors of APP; Captopril, Enalapril, Lisinopril, Cilazopril and salts thereof as inhibitors of ACE; Postatin, Eurystatin A or B, Naprotected peptide aldehydes, preferably benzyloxycarbonyl-L-prolyl-L-prolinal or benzyloxcarbonyl-L-thioprolyl-L-thioprolinal, Naprotected amino acid (Xaa)-pyrrolidides or -thiazolidides (Xaa=α-amino acid, preferably L-alanine, L-valine, L-isoleucine) as well as the corresponding 2-cyanopyrrolidid or 2-cyanothiazolidid derivatives, substrate-analogous Na-protected peptide phosphonic acid diaryl esters or peptide diazo methyl ketones or peptide ammonium methyl ketones and salts thereof as inhibitors of POP (PEP). 11. Pharmaceutical preparation according to claim 7 comprising two or more of the inhibitors of DP IV or of enzymes having a DP IV-analogous enzyme activity, of APN or of enzymes having an APN-analogous enzyme activity, of ACE, of POP (PEP) and of XPNPEP2 in a spaced apart formulation in combination with per se known carrier, auxiliary and/or additive substances for a simultaneous or a directly consecutive administration with the aim of a combined action. 12. Pharmaceutical preparation according to claim 7 for a systemic use of the oral, transdermal, intravenous, subcutaneous, intracutaneous, intramusclar, rectal, vaginal, sublingual application, together with per se known carrier, auxiliary and/or additive substances. 13. Pharmaceutical preparation according to claim 7 for a topical use in the form of, for example, creams, ointments, pastes, gels, solutions, sprays, liposomes, shaked mixtures, hydrocolloid dressings, and other dermatological bases/vehicles including instillative applications. 14. Use of inhibitor combinations of claim 2 for a prevention and therapy of autoimmune diseases, preferably of Rheumatoid Arthritis, Lupus Erythematodes, Multiple Sclerosis, IDDM, Morbus Crohn, Colitis Ulcerosa, Psoriasis, Neurodermitis, Glomerulonephritis, Interstitial Nephritis, Vasculitis, autoimmune diseases of the thyroid gland or autoimmune hemolytic anemia as well as of other chronic diseases with an inflammatory genesis as, for example, allergies and arteriosclerosis. 15. Use of inhibitor combinations of claim 2 for a suppression of transplant rejection and for a therapy of tumor diseases. 16. Use of inhibitor combinations of claim 3 for a prevention and therapy of autoimmune diseases, preferably of Rheumatoid Arthritis, Lupus Erythematodes, Multiple Sclerosis, IDDM, Morbus Crohn, Colitis Ulcerosa, Psoriasis, Neurodermitis, Glomerulonephritis, Interstitial Nephritis, Vasculitis, autoimmune diseases of the thyroid gland or autoimmune hemolytic anemia as well as of other chronic diseases with an inflammatory genesis as, for example, allergies and arteriosclerosis. 17. Use of inhibitor combinations of claim 3 for a suppression of transplant rejection and for a therapy of tumor diseases. 18. Use of inhibitor combinations of claim 4 for a prevention and therapy of autoimmune diseases, preferably of Rheumatoid Arthritis, Lupus Erythematodes, Multiple Sclerosis, IDDM, Morbus Crohn, Colitis Ulcerosa, Psoriasis, Neurodermitis, Glomerulonephritis, Interstitial Nephritis, Vasculitis, autoimmune diseases of the thyroid gland or autoimmune hemolytic anemia as well as of other chronic diseases with an inflammatory genesis as, for example, allergies and arteriosclerosis. 19. Use of inhibitor combinations of claim 4 for a suppression of transplant rejection and for a therapy of tumor diseases. 20. Pharmaceutical preparation according to claim 8 comprising two or more of the inhibitors of DP IV or of enzymes having a DP IV-analogous enzyme activity, of APN or of enzymes having an APN-analogous enzyme activity, of ACE, of POP (PEP) and of XPNPEP2 in a spaced apart formulation in combination with per se known carrier, auxiliary and/or additive substances for a simultaneous or a directly consecutive administration with the aim of a combined action. 21. Pharmaceutical preparation according to claim 8 for a systemic use for the oral, transdermal, intravenous, subcutaneous, intracutaneous, intramuscular, rectal, vaginal, sublingual application, together with per se known carrier, auxiliary and/or additive substances. 22. Pharmaceutical preparation according to claim 8 for a topical use in the form of, for example, creams, ointments, pastes, gels, solutions, sprays, liposomes, shaked mixtures, hydrocolloid dressings, and other dermatological bases/vehicles including instillative applications. 23. Pharmaceutical preparation according to claim 9 comprising two or more of the inhibitors of DP IV or of enzymes having a DP IV-analogous enzyme activity, of APN or of enzymes having an APN-analogous enzyme activity, of ACE, of POP (PEP) and of XPNPEP2 in a spaced apart formulation in combination with per se known carrier, auxiliary and/or additive substances for a simultaneous or a directly consecutive administration with the aim of a combined action. 24. Pharmaceutical preparation according to claim 9 for a systemic use for the oral, transdermal, intravenous, subcutaneous, intracutaneous, intramuscular, rectal, vaginal, sublingual application, together with per se known carrier, auxiliary and/or additive substances. 25. Pharmaceutical preparation according to claim 9 for a topical use in the form of, for example, creams, ointments, pastes, gels, solutions, sprays, liposomes, shaked mixtures, hydrocolloid dressings, and other dermatological bases/vehicles including instillative applications. 26. Pharmaceutical preparation according to claim 10 comprising two or more of the inhibitors of DP IV or of enzymes having a DP IV-analogous enzyme activity, of APN or of enzymes having an APN-analogous enzyme activity, of ACE, of POP (PEP) and of XPNPEP2 in a spaced apart formulation in combination with per se known carrier, auxiliary and/or additive substances for a simultaneous or a directly consecutive administration with the aim of a combined action. 27. Pharmaceutical preparation according to claim 10 for a systemic use for the oral, transdermal, intravenous, subcutaneous, intracutaneous, intramuscular, rectal, vaginal, sublingual application, together with per se known carrier, auxiliary and/or additive substances. 28. Pharmaceutical preparation according to claim 10 for a topical use in the form of, for example, creams, ointments, pastes, gels, solutions, sprays, liposomes, shaked mixtures, hydrocolloid dressings, and other dermatological bases/vehicles including instillative applications. 29. Pharmaceutical preparation according to claim 11 for a systemic use for the oral, transdermal, intravenous, subcutaneous, intracutaneous, intramuscular, rectal, virginal, sublingual application, together with per se known carrier, auxiliary and/or additive substances. 30. Pharmaceutical preparation according to claim 11 for a topical use in the form of, for example, creams, ointments, pastes, gels, solutions, sprays, liposomes, shaked mixtures, hydrocolloid dressings, and other dermatological bases/vehicles including instillative applications.

Induction heating of rail welds

An apparatus (10) and method for preheating welds uses a centered induction plate (12) having preferably a plurality of induction coils (30, 32) to impart the generation of heat in the materials to be welded (14), being interactively controlled by at least a temperature sensor (90, 92) and power supply control loop (16) so that even preheating can be obtained for a selected length of time given the parameters of the weld desired.

1. An induction heating system for preheating first and second workpieces to be welded together comprising: a tool having first and second opposing sides, and disposed in a gap defined by facing ends of the workpieces, the tool further including mechanical centering assemblies disposed adjacent the facing ends of the workpieces, wherein the mechanical centering assemblies position the tool longitudinally with respect to the workpieces for proper proximity to the workpieces; a first induction heating element affixed to the first side of the tool and disposed proximate the first and second workpieces; a first heating zone defined by a first temperature sensor positioned proximate the first workpiece and the first heating element; a second induction heating element affixed to the second side of the tool and disposed proximate the first and second workpieces; a second heating zone defined by a second temperature sensor positioned proximate the first workpiece and the second heating element; a controller coupled to the first and second temperature sensors, the controller activating the first and second heating elements independently to achieve a predetermined welding temperature in both first and second heating zones. 2. The induction heating system of claim 1, further comprising a first power supply coupled to the first heating element, and a second power supply coupled to the second heating element, wherein both first and second power supplies are coupled to the controller. 3. The induction heating system of claim 1, wherein the first and second induction heating elements are ferrite core heating elements. 4. The induction heating system of claim 1, wherein the temperature sensors comprise thermocouple temperature sensors. 5. The induction heating system of claim 1, wherein the temperature sensors comprise pyrometer temperature sensors. 6. The induction heating system of claim 1, wherein the controller provides a discernible indication when the workpieces have reached the predetermined welding temperature. 7. The induction heating system of claim 6, wherein the discernible indication is a visual indication. 8. The induction heating system of claim 1, wherein the controller provides positioning information to a positioning robot that adjusts relative position of the tool until the predetermined welding temperature is achieved. 9. The induction heating system of claim 1, wherein cooling fluid is provided proximate each of the heating elements. 10. The induction heating system of claim 9, wherein the controller controls cooling fluid activation. 11. The induction heating system of claim 1, wherein there are three or more heating elements. 12. The induction heating system of claim 1, wherein there are three or more heating zones. 13. The induction heating system of claim 1, wherein the first and second workpieces are railroad rails. 14. An induction heating system for preheating first and second workpieces to be welded together comprising: a tool disposed in a gap defined by facing ends of the workpieces, the tool further including mechanical centering assemblies disposed adjacent the facing ends of the work-pieces, wherein the mechanical centering assemblies position the tool longitudinally with respect to the workpieces for proper proximity to the workpieces; an induction heating element affixed to the tool and disposed proximate the first and second workpieces; a heating zone defined by a temperature sensor positioned proximate the first workpiece and the heating element; a controller coupled to the temperature sensor, the controller activating the heating element to achieve a predetermined welding temperature in the heating zone. 15. An induction heating system for preheating first and second workpieces to be welded together comprising: a tool disposed in a gap defined by facing ends of the workpieces, the tool further including mechanical centering assemblies disposed adjacent the facing ends of the workpieces, wherein the mechanical centering assemblies position the tool longitudinally with respect to the workpieces for proper proximity to the workpieces; a plurality of induction heating elements affixed to the tool and disposed proximate the first and second workpieces; as plurality of heating zones defined by multiple temperature sensors positioned proximate the first workpiece and the heating elements; a controller coupled to the temperature sensors, the controller activating the heating elements to achieve a predetermined welding temperature in the heating zones. 16. An induction heating system for preheating first and second workpieces to be welded together comprising: mounting means having first and second opposing sides, and disposed in a gap defined by facing ends of the workpieces, wherein the mounting means further further includes mechanical centering assemblies disposed adjacent the facing ends of the workpieces, wherein the mechanical centering assemblies position the mounting means longitudinally with respect to the workpieces for proper proximity to the workpieces; a first induction heating means affixed to the first side of the mounting means and disposed proximate the first and second workpieces; a first heating zone defined by a first temperature sensing means positioned proximate the first workpiece and the first heating means; a second induction heating means affixed to the second side of the mounting means and disposed proximate the first and second workpieces; a second heating zone defined by a second temperature sensing means positioned proximate the first workpiece and the second heating means; a controller means coupled to the first and second temperature sensing means, the controller means activating the first and second heating means independently to achieve a predetermined welding temperature in both first and second heating zones. 17. The induction heating system of claim 16, wherein the mounting means comprises a tool having first and second opposing sides. 18. The induction heating system of claim 16, further comprising a first power supply means coupled to the first heating means, and a second power supply means coupled to the second heating means, wherein both first and second power supplies means are coupled to the controller means. 19. The induction heating system of claim 16, wherein the first and second induction heating means comprise ferrite core heating elements. 20. The induction heating system of claim 16, wherein the temperature sensing means comprise thermocouple temperature sensors. 21. The induction heating system of claim 16, wherein the temperature sensing means comprise pyrometer temperature sensors. 22. The induction heating system of claim 16, wherein the controller means provides a discernible indication when the workpieces have reached the predetermined welding temperature. 23. The induction heating system of claim 22, wherein the discernible indication is a visual indication. 24. The induction heating system of claim 16, wherein the controller means provides positioning information to a positioning means that adjusts relative position of the tool until the predetermined welding temperature is achieved. 25. The induction heating system of claim 16, wherein cooling fluid is provided proximate each of the heating means. 26. The induction heating system of claim 25, wherein the controller means controls cooling fluid activation. 27. The induction heating system of claim 16, wherein there are three or more heating means. 28. The induction heating system of claim 16, wherein there are three or more heating zones. 29. The induction heating system of claim 16, wherein the first and second workpieces are railroad rails. 30. A method for preheating first and second workpieces to be welded together, the method comprising the steps of: (a) disposing, within a gap defined by facing ends of the workpieces, mounting means having first and second opposing sides; (b) providing mechanical centering assemblies disposed adjacent the facing ends of the workpieces, wherein the mechanical centering assemblies position the mounting means longitudinally with respect to the workpieces for proper proximity to the workpieces; (c) disposing a first induction heating means affixed to the first side of the mounting means proximate the first and second workpieces; (d) defining a first heating zone by positioning a first temperature sensing means proximate the first workpiece and the first heating means; (e) disposing a second induction heating means affixed to the second side of the mounting means proximate the first and second workpieces; (f) defining a second heating zone by positioning a second temperature sensing means proximate the first workpiece and the second heating means; and (g) activating the first and second heating means independently to achieve a predetermined welding temperature in both first and second heating zones.

<SOH> BACKGROUND OF THE INVENTION <EOH>1. Summary of Invention The preferred embodiment is adapted to use in welding railroad rails, however, other difficult to weld work pieces could be advantageously preheated with the invention. The descriptions herein of rail welds should be considered with this more expansive use in mind. Induction type heat allows for precise heating at ideal locations and can be used to control heat gradients. This control is possible though use of a feedback system, controller, coil arrangement and a positioning mechanism. This eliminates the human element, resulting in an automated high quality weld preheat method. The use of an automated electrically powered and computer controlled induction heating system using the induction heating coils and heating plate with sensitive temperature control and feedback interfaced within the power supply enables higher quality and more consistent welds of difficult to weld pieces such as railroad rails and similar high strength and complex shaped generally ferric items. One advantage in this regard is the ability to manipulate the heat gradient. Also included in advantages over prior art methods are the facts that no consumables are required, there is no need for gases or fuel on board or during transportation and cleanliness—in that there are no combustion byproducts. The invention provides consistent heat through a wide range of ambient temperatures. Another advantage is that of use in different rail geometry, rail chemistries and welding methods. In addition to heating, an analogous plate or array can be used to control cooling after welding. In the preferred and alternative embodiments, the invention envisions the use of independent, single or multiple coils and/or power units. Independent, single or multiple coils and/or power units enable the precise location of heating, subdivides locations of heating and provides flexibility in the control of heating areas. An added benefit of using an independent preheating unit, as compared to including the welder or portions of the welder's power supply or the like is that of efficiencies gains due to multi tasking during welding process. While preheating is occurring, the welder itself can be independently set up for welding operations, or, indeed, one rail may be welded while the adjacent rail is preheated, should the rail gaps be proximate the rail welder's cable runs. 2. Description of Related Art While preheating of metal pieces for welding as a general concept is well known, heretofore generally manual application of heat has been used. The use of items such as gas or other torches, gas burners or electrically powered devices. Field welding in the past commonly preheated with torches and gas burners. Such methods introduce human intervention positioning, timing or estimating heat input and temperature. Combustion variables including fuel, air, pressure, position and shape of a flame relative to rails, ignition steps, initial temperature of the workpieces and even weather contribute to imprecision in temperature control in the prior art. Resistance electrical devices have power and conductivity variables including both electrical and thermal limitations that also contribute to imprecision in temperature control.

<SOH> BRIEF DESCRIPTION OF THE DRAWINGS <EOH>FIG. 1 is a schematic view showing the layout of the components of the invention. FIG. 2 is an elevational view showing the induction plate of the invention. FIG. 2A is an elevational view showing an induction coil with ferrite core. FIG. 3 is a wiring diagram showing the control wiring of the invention. FIG. 4 is a wiring diagram showing the power supply wiring to control separate heating zones. FIG. 5 is a side elevational view showing the induction plate of the invention. detailed-description description="Detailed Description" end="lead"?

Carboxylate salts in heat-storage applications

The use of an alkali metal salt or alkaline earth metal salt or of a bring solution of a C1-C16 carboxylic acid, or a mixture thereof, as a medium for the storage and use of thermal energy. The salts or solution can be used in a heat exchange fluid or a lubricant or hydraulic fluid or soap.

1-13. (canceled) 14. A method for the storage and use of thermal energy comprising contacting one or a mixture of anhydrous alkali metal salts, alkali metal earth salts, amine salts or ammonium salts of a C1-C18 carboxylic acid or hydrated salts of a C1 to C7 carboxylic acid with a source of thermal energy. 15. The method of claim 1 further comprising contacting one or a mixture of anhydrous alkali metal salts, alkali metal earth salts, amine salts or ammonium salts of a C3-C18 carboxylic acid or hydrated salts of a C3 to C7 carboxylic acid with a source of thermal energy. 16. The method of claim 1 wherein said mixture comprises a salt of one or more C1-C2 carboxylic acids and a salt of one or more C3-C5 carboxylic acids. 17. The method of claim 1 wherein said mixture comprises a salt of one or more C1-C5 carboxylic acids and one or more C6-C18 carboxylic acids. 18. The method of claim 4 wherein said mixture comprises a salt of one or more C3-C5 carboxylic acids and one or more C6-C16 carboxylic acids. 19. The method of claim 1 wherein the temperature range of the thermal energy source is 20 to 180 degrees C. 20. The method of claim 1 further comprising contacting one or a mixture of anhydrous alkali metal salts, alkali metal earth salts, amine salts or ammonium salts of a C1 carboxylic acid with a source of thermal energy. 21. The method of claim 1 further comprising contacting one or a mixture of hydrated alkali metal salts, alkali metal earth salts, amine salts or ammonium salts of a C1 carboxylic acid with a source of thermal energy. 22. A method for improving the thermal properties of a fluid comprising dispersing in said fluid one or a mixture of anhydrous alkali metal salts, alkali metal earth salts, amine salts or ammonium salts of a C1-C18 carboxylic acid or hydrated salts of a C1 to C7 carboxylic acid with a source of thermal energy. 23. The method of claim 9 wherein the fluid is a water soluble alcohol freezing point depressant selected from the group consisting of ethylene glycol, propylene glycol, ethanol and methanol. 24. The method of claim 9 wherein the fluid is a lubricant or hydraulic fluid selected from the group consisting of mineral oil, synthetic oil, mineral soap, synthetic soap, mineral grease and synthetic grease.

<SOH> BACKGROUND OF THE INVENTION <EOH>Thermal energy originating from any energy source is reusable if it can be stored. Examples of reusable energy are excess heat from stationary and automotive internal combustion engines, heat generated by electrical motors and generators, process heat and condensation heat (e.g. in refineries and steam generation plants). Energy generated in peak load time can be managed and stored for later use. Examples are solar heating and electrical heating on low tariff hours. The problem of cold car engine start in wintertime is well known. Frost and damp on windscreen and windows, difficult engine start, cold in the passenger compartment. Car manufacturers are aware of this problem and make every possible effort to improve the driver's comfort under such circumstances. Electrical heating of windshield, rear windows, steering wheel and passenger seats are offered as comfort options. However these solutions put an extra burden on the vehicle's electrical power system. Engine manufacturers are looking for solutions that make preferably use of excess heat generated by the engine that can be controllably released to the environment. Heat-storage salts or functional fluids containing heat-storage salts may find new applications in emerging technologies. Heat-storage salts could for instance be applied to maintain fuel cells at constant temperatures. An aspect of this invention is that in automotive and heavy-duty engine applications, excess engine heat can be stored in carboxylic salts or in carboxylic salt solutions integrated into the engine heat-exchange system. The stored heat can be used to rapidly heat critical engine components, engine fluids and exhaust gas catalyst Heating of these critical components before engine start helps avoids the discomfort, high fuel consumption, high exhaust emissions and increased engine wear linked to cold engine start. The heat stored in carboxylic salts or in carboxylic salts solutions can also be used to heat the passenger compartment to improve driver and passenger comfort in cold climates.

Stabiliser for radiopharmaceuticals

The present invention provides an improved stabiliser for radiopharmaceuticals which inhibits impurities from being produced by two kinds of decomposition mechanisms and exhibits such an effect that the shelf life of a radiopharmaceutical after its preparation is prolonged as compared with conventional ones. The improvement comprises a combination of an amino-substituted aromatic carboxylic acid or its salt, ester or amide in combination with a diphosphonic acid or its salt.

1. A stabilised radiopharmaceutical composition which comprises: (i) a radiopharmaceutical which is susceptible to either reductive degradation or radiolysis; (ii) a first stabiliser for said radiopharmaceutical which includes an amino-substituted aromatic carboxylic acid or a salt, ester or amide thereof; (iii) a second stabiliser for said radiopharmaceutical which includes a diphosphonic acid or salt thereof, wherein the radiopharmaceutical is not a metal complex of the diphosphonic acid. 2. The composition of claim 1 wherein the amino-ubstituted aromatic carboxylic acid or its salt, ester or amide is selected from the group consisting of 2-aminobenzoic acid, 3-aminobenzoic acid, 4-aminobenzoic acid, 3, 5-diaminobenzoic acid, 4-amino-salicylic acid, and salts, esters and amides thereof. 3. The composition of claim 1, wherein the diphosphonic acid or its salt is selected from the group consisting of methylenediphosphonic acid, hydroxymethanediphosphonic acid, hydroxyethanediphosphonic acid, ethylenediaminetetraphosphonic acid, and salts thereof. 4. The composition of claim 1, wherein the amino-substituted aromatic carboxylic acid or salt, ester or amide thereof is sodium 4-aminobenzoate, and the diphosphonic acid is methylenediphosphonic acid. 5. The composition of claim 1, wherein the radiopharmaceutical includes a γ-ray emitter or a β-ray emitter. 6. The composition of claim 5, wherein the radiopharmaceutical includes Tc-99m, Re-186 or Re-188. 7. The composition of claim 1, wherein the radiopharmaceutical includes a metal complex of a radionuclide with a ligand. 8. The composition of claim 7, wherein the ligand is an amine oxime. 9. The composition of claim 8, wherein the amine oxime is selected from the group consisting of d,l-hexamethylpropyleneamine oxime and 4,9-diaza-3, 3, 10, 10-tetramethyldodecane-2, 11-dione dioxime. 10. A non-radioactive kit for the preparation of the stabilised radiopharmaceutical composition of claim 1 which comprises: (i) a first stabiliser which comprises an amino-substituted aromatic carboxylic acid or a salt, ester or amide thereof; and (ii) a second stabiliser which comprises a diphosphonic acid or salt thereof, wherein the radiopharmaceutical is not a metal complex of the diphosphonic acid. 11. The kit of claim 10, which includes 0.01 to 10 mg of an amino-substituted aromatic carboxylic acid or its salt, ester or amide, and 0.01 to 1 mg of a diphosphonic acid or its salt. 12. The kit of claim 10, further comprising a tin (II) reductant, wherein the molar ratio of the diphosphonic acid or its salt to tin (II) is in the range 1 to 10. 13. The kit of claim 10, further comprising an amine oxime ligand. 14. The kit of claim 13 wherein the amine oxime is selected from the group consisting of d, l-hexamethylpropyleneamine oxime and 4,9-diaza-3, 3, 10, 10-tetramethyldodecane-2, 11-dione dioxime. 15. The kit of claim 10, wherein the first and second stabilzers are lyophilised. 16. In a method of preparing a radiopharmaceutical composition which includes adding stabilizers to a radiopharmaceutical, wherein the radiopharmaceutical is not a metal complex of diphosphonic acid, the improvement comprising using a diphosphonic acid or a salt thereof as a stabiliser. 17. Cancelled 18. The method of claim 16, wherein the radiopharmaceutical comprises Tc-99m. 19. The method of claim 16, wherein the diphosphonic acid is selected from the group consisting of methylenediphosphonic acid, hydroxymethanediphosphonic acid, hydroxyethanediphosphonic acid, ethylenediaminetetraphosphonic acid, and salts thereof.

<SOH> BACKGROUND TO THE INVENTION <EOH>Some radiopharmaceuticals undergo decomposition due to either radiolysis or redox reactions, and hence exhibit undesirable instability. Non-radioactive kits for the preparation of radiopharmaceuticals, especially Tc-99m radiopharmaceuticals, may suffer from two types of instability: (i) shelf-life instability of the non-radioactive composition over time, (ii) instability of the radiopharmaceutical post-formation. In the case of Tc-99m, the latter is referred to as post-reconstitution instability. U.S. Pat. No. 4,451,451 discloses that para-aminobenzoic acid (pABA) and analogues are useful stabilisers for technetium non-radioactive kits, including kits for the preparation of 99m Tc-complexes of diphosphonic acids. Tc-99m-hexamethylpropyleneamine oxime (referred to hereinafter as 99m Tc-HMPAO), is a radiopharmaceutical commercially available as a regional cerebral blood flow imaging agent. 99m Tc-HMPAO is particularly unstable with respect to post-reconstitution stability. 99m Tc-HMPAO is usually prepared from a lyophilised, non-radioactive kit which contains HMPAO and stannous ion. The function of the stannous ion is to reduce the 99m Tc-pertechnetate ( 99m TcO 4 − ), ie. technetium in oxidation state Tc(VII), to the Tc(V) oxidation state of the 99m Tc-HMPAO metal complex. The radiochemical purity (Rcp) of 99m Tc-HMPAO one hour after Tc-99m labelling is only about 80%, so that it must be used within 30 minutes of 99m Tc labelling. After Tc-99m radiolabelling, the Rcp of 99m Tc-HMPAO decreases with time due to the growth of three different radioactive impurities, namely: a hydrophilic secondary 99m Tc complex of unknown structure derived from 99m Tc-HMPAO, 99m Tc-pertechnetate ( 99m TcO 4 − ) and reduced-hydrolysed-technetium [ 99m Tc]. Of these impurities, both the secondary complex and 99m Tc-pertechnetate are decomposition products of 99m Tc-HMPAO; however, it is reported that the decomposition mechanisms are different (J. Nucl. Med. 29, 1568-1576, 1988). The secondary complex is believed to be produced when the lipophilic 99m Tc-HMPAO complex is exposed to excess unoxidised tin(II) (ie. stannous) remaining from the pertechnetate reduction step. On the other hand, the 99m Tc-pertechnetate impurity is produced when 99m Tc-HMPAO and the secondary complex are oxidised by the free radicals produced in solution by the action of radiation, ie. radiolysis of the solvent. Accordingly, in order to inhibit the production of both the 99m Tc-pertechnetate and secondary complex impurities, the addition of stabilisers has been disclosed. Thus, Nucl. Med. Biol. 7, 675-680 (1989); Eur. J. Nucl. Med. 16, 541 (1990); Eur. J. Nucl. Med. 20, 661-666 (1993) and Eur. J. Nucl. Med. 22, 1163-1172 (1995) all report attempts to stabilise 99m Tc-HMPAO by the addition of either: gentisic acid, sodium decahydroxypyrophosphate, methylene blue, cobalt chloride or the like. In particular, the post-radiolabelling addition of methylene blue improves the Rcp of 99m Tc-HMPAO to at least 80% at 4 hours post reconstitution. Similarly, the post-radiolabelling addition of cobalt chloride has been found to improve the Rcp of 99m Tc-HMPAO at 6 hours post reconstitution to at least 80%. The stabilisation mechanisms of 99m Tc-HMPAO by methylene blue and cobalt chloride are believed to be essentially the same. Both are in redox equilibrium in solution, and oxidise excess tin(II), thus stabilising 99m Tc-HMPAO. However, when the reducing tin(II) and methylene blue or cobalt chloride coexist in solution before the Tc-99m radiolabelling step, the tin(II) reductant is completely oxidised, so that the Tc-99m labelling becomes impossible because there is no longer any reducing agent present to reduce the Tc(VII) 99m Tc-blue or cobalt chloride is used as a stabiliser for 99m Tc-HMPAO, it must be added after the Tc-99m radiolabelling step and cannot be pre-mixed with the ligand (HMPAO) and 99m Tc-pertechnetate. Accordingly, any kit for the preparation of 99m Tc-HMPAO employing such stabilisers, must be composed of two vials (referred to hereinafter as a 2-vial kit). One vial is a freeze-dried vial containing the HMPAO ligand together with the tin(II) reductant and other excipients. The other is a vial containing the stabiliser (methylene blue or cobalt chloride). Thus, the most successful prior art methods of stabilising 99m Tc-HMPAO to date all require the use of 2-vial kits. When the kit for preparing 99m Tc-HMPAO is a 2-vial kit, the radiolabelling operation is more complicated than for a single vial kit and comprises two steps: (1) 99m Tc-pertechnetate solution is added to the vial containing the HMPAO ligand and the resulting mixture is mixed by shaking; (2) a stabiliser solution from the second vial (eg. methylene blue or cobalt chloride) is added to the reconstituted mixture from step (1) in the first vial. It is necessary that the time between the first and second steps is controlled so as to be as close as possible to two minutes. Too short a time, and 99m Tc-HMPAO complex formation may be incomplete and hence addition of the stabiliser may adversely affect the Rcp by oxidising the stannous ion before the reduction of the pertechnetate starting material is complete. Too long a time, and the stabilising effect is delayed. In such a procedure it is also necessary that care be taken with respect to the amounts of the solutions added. The operator must also take due care to ensure that the vials are not inadvertently mixed up at any stage. In addition, there is an increased risk of radiation dose to the operator due to the increased number of manipulations. Moreover, when methylene blue is added to 99m Tc-HMPAO, a precipitate is produced, so that a filtration step becomes necessary, and thus the procedure becomes more complicated. There is therefore a need for a single vial kit for the preparation of 99m Tc-HMPAO which has both shelf-life and post-radiolabelling stability. The present invention provides a kit which solves this problem, and is straightforward to use. The Present Invention. The present invention relates to a stabiliser for radiopharmaceuticals which comprises a combination of an amino-substituted aromatic carboxylic acid or a salt, ester or amide thereof, with a diphosphonic acid or a salt thereof with the proviso that the radiopharmaceutical is not a metal complex of the diphosphonic acid.

Refrigeration unit

The refrigerating unit can be operated by means of a thermal solar system as energy source. Therein it is provided that the refrigerating unit is formed as a diffusion-absorption refrigerating unit. The refrigerating unit has an expeller, a triple heat exchanger, a condenser, an evaporator, a gas heat exchanger, an absorber, and a fuel reservoir which are actively connected to form a closed fuel circuit with one another.

1-12. Cancelled. 12. A refrigerating unit, which is formed as a diffusion-absorption refrigerating unit, comprising: a closed fuel circuit having a triple heat exchanger for preheating a solution which is high in fuel and precooling a solution which is low in fuel, and for rectification of a mixture of fuel and water vapor. 13. The refrigerating unit according to claim 12, further comprising a thermal solar system actively connected to an expeller formed as a gas bubble pump for the desorption and vaporization of a fuel contained in a solution. 14. The refrigerating unit according to claim 13, wherein the the expeller has at least one rising tube which on the inlet side is provided with a ribbing promoting the vaporization of the fuel contained in the rising tube. 15. The refrigerating unit according to claim 13, wherein the the expeller has at least one rising tube which on the inlet side is provided with a ribbing promoting the vaporization of the fuel contained in the rising tube. 16. The refrigerating unit according to claim 13, wherein the expeller has a plurality of parallel rising tubes spaced at a distance from one another for the conveyance of a solution which is high in fuel, where a parallel recycling line for the conveyance of the solution which is low in fuel is disposed between at least two rising tubes. 17. The refrigerating unit according to claim 13, further comprising a rising tube and wherein the expeller is actively connected, on an inlet side with respect to the rising tube, to a supply line, coming from the triple heat exchanger, for the conveyance of the solution which is high in fuel, on the outlet side with respect to the rising tube, to a fuel vapor supply line leading to the triple heat exchanger, and, on the outlet side with respect to the recycling line, to a supply line, leading to the triple heat exchanger, for the conveyance of the solution which is low in fuel. 18. The refrigerating unit according to claim 17, wherein the fuel vapor supply line leads essentially coaxially through the recycling line. 19. The refrigerating unit according to claim 13, further comprising a rising tube having an inner structure promoting the formation of bubbles. 20. The refrigerating unit according to claim 12, wherein the fuel is ammonia (NH3) and the solution a mixture of ammonia and water (NH3/H2O). 21. The refrigerating unit according to claim 13, wherein the thermal solar system is actively connected over an entire longitudinal extension of the rising tube of the expeller. 22. The refrigerating unit according to claim 13, further comprising at least one of an absorber and an evaportator having conveyance lines, wherein the conveyance lines have upper free ends with a corresponding capillary sleeve 23. The refrigeration unit of claim 12, where the closed fuel circuit comprises an expeller condenser, an evaporator, a gas heat exchanger, an absorber, and a fuel reservoir which are actively connected to one another to form a closed fuel circuit. 24. The refrigerating unit according to claim 23, further comprising a thermal solar system actively connected to an expeller formed as a gas bubble pump for the desorption and vaporization of a fuel contained in a solution. 25. The refrigerating unit according to claim 23, wherein the the expeller has at least one rising tube which on the inlet side is provided with a ribbing promoting the vaporization of the fuel contained in the rising tube. 26. The refrigerating unit according to claim 23, wherein the the expeller has at leapt one rising tube which on the inlet side is provided with a ribbing promoting the vaporization of the fuel contained in the rising tube. 27. The refrigerating unit according to claim 12, wherein the expeller has a plurality of parallel rising tubes spaced at a distance from one another for the conveyance of a solution which is high in fuel, where a parallel recycling line for the conveyance of the solution which is low in fuel is disposed between at least two rising tubes. 28. The refrigerating unit according to claim 23, further comprising a rising tube and wherein the expeller is actively connected, on an inlet side with respect to the rising tube, to a supply line, coming from the triple heat exchanger, for the conveyance of the solution which is high in fuel, on the outlet side with respect to the rising tube, to a fuel vapor supply line leading to the triple heat exchanger, and, on the outlet side with respect to the recycling line, to a supply line, leading to the triple heat exchanger, for the conveyance of the solution which is low in fuel. 29. The refrigerating unit according to claim 28, wherein the fuel vapor supply line leads essentially coaxially through the recycling line. 30. The refrigerating unit according to claim 23, further comprising a rising tube having an inner structure promoting the formation of bubbles. 31. The refrigerating unit according to claim 23, wherein the fuel is ammonia (NH3) and the solution a mixture of ammonia and water (NH3/H2O). 32. The refrigerating unit according to claim 23, wherein the thermal solar system is actively connected over an entire longitudinal extension of the rising tube of the expeller. 33. The refrigerating unit according to claim 23, further comprising at least one of an absorber and an evaportator having conveyance lines, wherein the conveyance lines have upper free ends with a corresponding capillary sleeve 34. A refrigerating unit, which is formed as a diffusion-absorption refrigerating unit and can be operated by means of a thermal solar system as energy source, with an expeller, a triple heat exchanger, a condenser, an evaporator, a gas heat exchanger, an absorber, and a fuel reservoir which are actively connected to one another to form a closed fuel circuit, wherein the refrigerating unit has, in the fuel circuit, a triple heat exchanger for preheating a solution which is high in fuel and precooling a solution which is low in fuel, and for the rectification of a mixture of fuel and water vapor. 35. A method for refrigerating a unit, the method comprising: forming a fuel circuit having a triple head exchanger; and preheating a solution which is high in fuel, precooling a solution which is low in fuel, and rectifying a mixture of fuel and water vapor.

<SOH> BACKGROUND <EOH>The invention relates to a refrigerating unit, which can be operated by means of a thermal solar system as energy source, according to the preamble of claim 1 . Refrigerating units of the type addressed here, such as, for example, absorption refrigerating units or compression refrigerating units, are known. For solar operation, these absorption refrigerating units disadvantageously require a high outlay with regard to control technology and investment, in particular with regard to a continuous reheating or heat storage. In view of this, absorption refrigerating units are limited to a relatively narrow temperature and flow range. Also the known, photovoltaically operated compression refrigerating units are characterized by a high outlay with regard to investment and have relatively low efficiency. These known refrigerating units are not suited to problem-free and economical operation with thermal solar systems, in particular in the average capacity range.

<SOH> SUMMARY OF THE INVENTION <EOH>It is the objective of the invention to provide a refrigerating unit of the type stated initially which can be operated in a reliable, effective, and operation-friendly manner by means of a thermal solar system, in particular in the lower to average refrigerating capacity range. For the realization of this objective, a refrigerating unit with the features of claim 1 is proposed which is distinguished by the fact that the refrigerating unit is formed as a diffusion-absorption refrigerating unit. A diffusion-absorption refrigerating unit can be operated in a particularly reliable and efficient manner by means of a thermal solar system as an energy source. This behavior, which is favorable to operation, can also be achieved in a smaller to average refrigerating capacity range. A diffusion-absorption refrigerating unit is advantageously suitable to be operated by means of various energy sources. Among these, a thermal solar system as well as another heat transfer medium circuit, e.g. from a heat recovery process, can be used for the alternative or enhancing energizing of the refrigerating unit. A diffusion-absorption refrigerating unit is thus advantageously suitable, in a manner which is flexible and favorable for operation, to be energized with thermal energy by means of a thermal solar system as well as, if needed or desired, additional, different energy sources. The refrigerating unit advantageously has an expeller, a triple heat exchanger, a condenser, an evaporator, a gas heat exchanger, an absorber, and a fuel reservoir, which are actively connected to one another to form a closed fuel circuit. The refrigerating unit characterized by a closed fuel circuit can be formed as a hermetically closed, compact unit which is distinguished by an advantageous independence of site. It is operated merely by means of thermal energy and thus advantageously needs no electrical power supply. Since no components working mechanically within the refrigerating unit, such as, for example, pumps, are required to operate the refrigerating unit, the unit is maintenance-friendly, relatively favorable from the standpoint of cost, and can be operated, at least nearly, without noise. Furthermore, it is possible to develop the refrigerating unit so that the mounting of several refrigerating units in parallel can be realized in a relatively simple manner. Preferably, the thermal solar system can be actively connected to an expeller formed as a gas bubble pump for the desorption and vaporization of a fuel contained in a solution. A gas bubble pump is particularly suitable for desorbing and vaporizing, in a manner which is effective and favorable for operation, a fuel contained in a solution such as, for example, ammonia (NH3) in an ammonia-rich solution. Furthermore, a gas bubble pump permits an efficient heat transfer accomplished by means of a thermal energy source, which is a prerequisite for reliable and effective desorption and vaporization of the fuel (ammonia). According to a preferred form of embodiment, the expeller has at least one rising tube which on the inlet side is provided with a ribbing promoting the vaporization of the fuel contained in the rising tube. The rising tube of the expeller formed as a gas bubble pump contains a solution which is high in fuel and which, due to the vaporization of the fuel in the rising tube, experiences an increase in volume so that it assumes an operating volume which is a multiple of the original volume value, i.e. of the still not vaporized fuel. Due to this increase in volume, the level of the vaporous fuel, or of the fuel mixture in the rising tube, rises to a corresponding operational conveyance height. If this operational conveyance height exceeds the length of the rising tube due to a corresponding increase in volume of the fuel mixture or a vaporization of the fuel, the conveyance of the vaporous fuel, or a remaining solution which is low in fuel, to additional functional units of the refrigerating unit is started, said functional units being actively connected to one another in the form of a closed fuel circuit so that the refrigerating unit is activated at this moment in its operation. In so doing, an effective heat transfer from a thermal energy source (solar system) into the expeller or into the rising tube serves for a reliable and rapid activation of the refrigerating unit. This is guaranteed by means of a suitable ribbing of the rising tube whereby an enlargement of the heat transfer surface to improve the heat transfer into the rising tube is obtained. Instead of ribbing, deflecting plates can also be provided. Advantageously, the expeller has a plurality of parallel rising tubes spaced at a distance from one another for the conveyance of a solution which is high in fuel, where a parallel recycling line for the conveyance of the solution which is low in fuel is disposed between at least two rising tubes. This makes possible an expeller formation which is compact and has the form of a bundle of tubes, where advantageously the rising tubes disposed externally can be energized with thermal energy, in a manner favorable to efficiency and operation, by means of a thermal energy source (solar system) while the interposed, parallel recycling line, through which a heated solution which is low in fuel is conveyed, makes possible an additional heating of the outer rising tubes, or counteracts a cooling of the same. The amount of heat radiating, and thus not actively utilized, is reduced in an effective manner due to a compact arrangement of the outer rising tubes as a bundle of tubes and a centrally disposed recycling line. Preferably, the expeller is actively connected, on the inlet side with respect to the rising tube, to a supply line coming from the triple heat exchanger for the conveyance of the solution which is high in fuel, on the outlet side with respect to the rising tube, to a fuel vapor supply line leading to the triple heat exchanger, and, on the outlet side with respect to the recycling line, to a supply line leading to the triple heat exchanger for the conveyance of the solution which is low in fuel. The expeller preferably having a compact bundle of tubes is thus actively connected, on the inlet side as well as on the outlet side, to the triple heat exchanger which in turn is actively connected to additional functional units of the refrigerating unit to form a closed fuel circuit. In so doing, a solution which is high in fuel, a solution which is low in fuel, and fuel vapor are conveyed in the expeller after activation of the refrigerating unit. Advantageously, the fuel vapor supply line leads essentially coaxially through the recycling line. An expeller of this type is formed particularly compactly and is distinguished by a particularly effective and reliable desorption and vaporization of a fuel in a solution which is high in fuel, said solution being contained in the respective rising tube, since an undesirable radiation of heat from the expeller into the environment can be reduced to a significant extent due to its compact arrangement as a bundle of tubes. Advantageously, the rising tube has an inner structure promoting the formation of bubbles. An inner structure of this type in the rising tube can, for example, be achieved by means of a coating and/or a roughening of the inner surface and serves for the desired bubble formation in a solution which is high in fuel in a definite area in the rising tube due desired turbulence of the same solution, said turbulence being caused by the inner structure. The expeller formed as a gas bubble pump is suitable, with the use of rising tubes of this type, for a particularly effective desorption, vaporization, and conveyance of fuel contained in a solution which is high in fuel and for the conveyance of a solution which is low in fuel. Advantageously, the fuel is ammonia (NH3) and the solution a mixture of ammonia and water (NH3/H 2 O). NH3 as fuel and a mixture of NH3 and H 2 O as solution are particularly suited to an effective operation of the refrigerating unit according to the invention. Along with this, an auxiliary gas, used, if needed or desired, for the operation of the refrigerating unit, can be helium (He). According to a preferred form of embodiment, the thermal solar system is actively connected over the entire longitudinal extension of the rising tube of the expeller to the same. By means of heating of the rising tube over its entire longitudinal extension, it is avoided in a reliable manner that, due to an undesired cooling in an unheated area of the rising tube, vaporous NH3, already expelled, condenses and thus cancels an intended increase in volume of the fuel contained in the rising tube, where the increase in volume would lead to the starting of the conveyance of the fuel, or the solution which is low in fuel. In this, the rising tubes are preferably each provided on the inlet side with a ribbing for the heating of the triple heat exchanger, of a fuel reservoir, and of a supply line leading to the condenser (vapor line). Additional advantageous developments of the invention follow from the description.

Acetyl glucosaminyl inositol deacetylase, a mycothiol biosynthetic enzyme, and methods of use

The present invention provides a family of bacterial acetyl glucosaminyl inositol deacetylases (MshB) with deacetylase activity against acyl glucosaminyl inositol and which play a key role in mycothiol biosynthesis. The invention deacetylases are characterized by a conserved 100 amino acid N-terminal region and three highly conserved histidine-containing regions and by having deacetylase activity as well as amide hydrolase activity. The invention further provides methods for using the invention deacetylases in drug screening assays to determine compounds that inhibit activity. The invention provides for treatment of actino-mycete infections in mammals using antibiotics that inhibit production or activity of MshB and thereby reduce the production of mycothiol and the virulence of the infecting bacteria.

1. A purified acetyl glucosaminyl inositol deacetylase, characterized as having: a) an N-terminal region with an amino acid sequence with 40% or more sequence identity to SEQ ID NO:2 and conservative variations thereof, b) three domains of conservation, wherein two of the domains contain conserved histidine residues, and c) deacetylase activity against acetyl glucosaminyl inositol. 2. The purified deacetylase of claim 1, wherein the deacetylase hydrolyzes a C2-amide bond of the glucosaminyl inositol moiety. 3. The purified deacetylase of claim 1, wherein the acetyl glucosaminyl inositol is a precursor of mycothiol. 4. The purified deacetylase of claim 1, wherein the acetyl glucoasaminyl inositol is N-acetyl-1-D-myo-inosityl-2-amino-2-deoxy-α-D-glucopyranoside (GlcNAc-Ins). 5. The purified deacetylase of claim 1, wherein the three domains have amino acid sequences selected from the group consisting of SEQ ID NOS: 3, 4, 5, 6, conservative variations thereof, and any combination of two or more thereof. 6. The purified deacetylase of claim 1, wherein the deacetylase is derived from an actinomycetes. 7. The purified deacetylase of claim 6, wherein the deacetylase is derived from M. smegmatis. 8. The purified deacetylase of claim 6, wherein the deacetylase is derived from M. tuberculosis. 9. The purified deacetylase of claim 6, wherein the deacetylase is derived from M. leprae. 10. The purified deacetylase of claim 6, wherein the deacetylase is derived from M. bovis. 11. The purified deacetylase of claim 6, wherein the deacetylase is derived from M. smegmatis, M. tuberculosis, M. leprae, M. bovis, M. intracellulare, M. africanum, M. marinarum. M. chelonai, Corynebacterium diphtheriae, Actinomyces israelii, M. avium complex (MAC), M. ulcerans, M. abscessus, or M. scrofulaceum. 12. The purified deacetylase of claim 6, wherein the bacterium is selected from the group consisting of Streptomyces lincolnensis, Amycolatopsis mediterranei, Amycolatopsis orientalis, Streptomyces lavendulae, Streptomyces coelicolor, Streptomyces rochei and Saccharopolyspora erythraea. 13. The purified deacetylase of claim 1, wherein the deacetylase has amidase activity against acyl glucosaminyl inositols. 14. The purified deacetylase of claim 1, wherein the deacetylase has an amino acid sequence as set forth in SEQ ID NO:1 15. The purified deacetylase of claim 1, wherein the deacetylase is encoded by a polynucleotide comprising a nucleic acid sequence as set forth in SEQ ID NO:7. 16. A purified acetyl glucosaminyl inositol deacetylase, characterized as having: a) an N-terminal region with an amino acid sequence with at least 40% sequence identity to SEQ ID NO:2, b) one or more domains of conservation containing conserved metal chelating residues, and c) deacetylase activity against acetyl glucosaminyl inositol in the presence of metal ion. 17. The purified deacetylase of claim 16, wherein the metal ion is selected from the group consisting of Mn2+ and Ni2, Cd2+, Co2+, and Zn2+. 18. The purified deacetylase of claim 17 wherein the metal ion is Zn2+. 19. The purified deacetylase of claim 16, wherein the metal chelating residues are selected from the group consisting of histidine, aspartic acid and glutamic acid, and combinations thereof. 20. An antibody, or functional fragment thereof, that binds specifically to an deacetylase of claim 1. 21. An isolated polynucleotide that encodes a deacetylase of claim 1. 22. A vector containing a polynucleotide that encodes an deacetylase of claim 1. 23. A cell transformed with a vector of claim 22. 24. A method for identifying an inhibitor of acetyl glucosaminyl inositol deacetylase, said method comprising: a) contacting a candidate compound with a deacetylase of claim 1 in the presence of an acyl glucosaminyl inositol under suitable conditions and b) determining the presence or absence of breakdown products of the acyl glucosaminyl inositol indicative of deacetylase activity or amidase activity, wherein the substantial absence of the deacetylase activity or the amidase activity is indicative of a candidate compound that inhibits activity of the deacetylase. 25. The method of claim 24, wherein the deacetylase is an acetyl glucosaminyl inositol deacetylase. 26. The method of claim 24, wherein a breakdown product is a free amine. 27. The method of claim 26, wherein the free amine is GlcN-Ins. 28. The method of claim 24, wherein a breakdown product is acetate 29. The method of claim 24, wherein a breakdown product is a derivative of cysteine. 30. The method of claim 24, wherein the acyl glucosaminyl inositol is N-acetyl-1-D-myo-inosityl-2-amino-2-deoxy-α-D-glucopyranoside. 31. The method of claim 24, wherein the acyl glucosaminyl inositol is an S-conjugate of mycothiol. 32. The method of claim 31, wherein the S-conjugate of mycothiol is the monobromobimane derivative of mycothiol (MSmB). 33. The method of claim 24, wherein the three domains in the deacetylase that contain conserved histidine residues have amino acid sequences selected from SEQ ID NOs: 3, 5, 6 and any combination of two or more thereof. 34. The method of claim 24, wherein the deacetylase is produced in an actinomycete. 35. The method of claim 34, wherein the actinomycete is M. smegmatis. 36. The method of claim 34, wherein the actinomycete is M. tuberculosis. 37. The method of claim 34, wherein the actinomycete is M. leprae. 38. The method of claim 34, wherein the actinomycete is M. bovis. 39. The method of claim 34, wherein the actinomycete is M. intracellulare, M. africanum, M. marinarum, M. chelonai, Corynebacterium diphtheriae, Actinomyces israelii, M. avium complex (MAC), M. ulcerans, M. abscessus, or M. scrofulaceum. 40. The method of claim 24, wherein candidate compound is a polypeptide, polynucleotide or small molecule. 41. A high throughput screening method for identifying inhibitors of the deacetylase of claim 1, said method comprising: a) contacting each of a plurality of candidate compounds with a deacetylase of claim 1 in the presence of acyl glucosaminyl inositol under suitable conditions to form a plurality of reaction mixtures and b) determining the presence or absence of binding to the reaction mixtures of a detectable marker that binds to free amine, wherein the substantial absence of binding of the marker to a reaction mixture is indicative of a candidate compound that inhibits activity of the deacetylase. 42. The method of claim 41, wherein the plurality of reaction mixtures are formed in the wells of a microtiter plate. 43. The method of claim 41, wherein the detectable marker is colorometric or fluorometric. 44. The method of claim 43 wherein the fluorometric marker is fluorescamine or 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate. 45. An inhibitor of the deacetylase of claim 1 wherein the inhibitor is derived from GlcN-Ins by replacing the amino group therein with a moiety that chelates Zn2+, or otherwise binds the enzyme active site of the deacetylase. 46. The inhibitor of claim 45 wherein the inhibitor additionally chelates one or more metal ions metal ions selected from the group consisting of Mn2+ and Ni2, Cd2+, and Co2+. 47. A derivative of GlcN-Ins, wherein the derivative contains a reactive residue attached to the amino group that promotes oxidative stress so as to be selectively toxic to a mycothiol-producing actinomycete by being concentrated in the actinomycete. 48. The derivative of claim 47, wherein the reactive residue is a nitroso residue. 49. The derivative of claim 47, wherein the reactive residue is a nitroalkyl residue comprising 1 to 3 carbon atoms. 50. The derivative of claim 47, wherein the reactive residue is a S-nitrosomercaptoalkyl residue comprising 2 to 4 carbon atoms. 51. The derivative of claim 47, wherein the reactive residue is a peroxyalkyl residue comprising 2 to 4 carbon atoms. 52. A live mutant actinomycete, whose genome comprises a disruption in an endogenous acetyl glucosaminyl inositol deacetylase gene, wherein said disruption prevents function of an endogenous acetyl glucosaminyl inositol deacetylase while cell surface proteins and lipids are substantially unaffected, and wherein said disruption results in said mutant actinomycetes exhibiting transient survival in mammalian white blood cells for an immune response-raising period of time. 53. The live mutant actinomycete of claim 52, wherein the period of time is from 1 to 30 days. 54. The live mutant actinomycete of claim 52, wherein the survival of the mutant actinomycetes in mammalian white blood cells does not exceed 30 days. 55. The live mutant actinomycete of claim 52, wherein the mutant actinomycete is derived from a pathogen selected from the group consisting of M. smegmatis, M. tuberculosis, M. leprae, M. bovis, M. intracellulare, M. africanum, M. marinarum, M. chelonai, Corynebacterium diphtheria, Actinomyces israelii, M. avium complex (MAC), M. ulcerans, M. abscessus, and M. scrofulaceum. 56. The live mutant actinomycete of claim 52, wherein the mutant actinomycetes. is derived from M. smegmatis. 57. The live mutant actinomycete of claim 52, wherein the mutant actinomycetes is derived from M. tuberculosis. 58. The live mutant actinomycete of claim 52, wherein the mutant actinomycetes is derived from M. leprae. 59. The live mutant actinomycete of claim 52, wherein the mutant actinomycetes is derived from M. bovis and said mammal is bovine. 60. A method for decreasing the virulence of a pathogenic acetyl glucosaminyl inositol deacetylase-producing bacterium in mammalian cells, said method comprising: introducing into the bacterium an inhibitor of acetyl glucosaminyl inositol deacetylase activity, wherein the intracellular presence of the inhibitor decreases activity of the deacetylase, thereby decreasing mycothiol biosynthesis by the bacterium as compared with untreated control bacterium. 61. The method of claim 60, wherein the inhibitor inhibits intracellular production of the deacetylase. 62. The method of claim 60, wherein the inhibitor inhibits intracellular deacetylase activity of the deacetylase. 63. The method of claim 60, wherein the introducing comprises culturing the bacterium in the presence of the inhibitor. 64. The method of claim 60, wherein the inhibitor is an anti-sense oligonucleotide complementary to a target region in a messenger RNA that encodes a polypeptide having an amino acid sequence segment with 40% or more sequence identity to the amino acid sequence of SEQ ID NO:2 or at least one amino acid sequence selected from the group consisting of SEQ ID NOS: 3, 4, 5, 6 and conservative variations thereof. 65. The method of claim 60, wherein the inhibitor is an anti-sense oligonucleotide that hybridizes under intracellular conditions with a messenger RNA that encodes a polypeptide having an N-terminal amino acid sequence as set forth in SEQ ID NO:2. 66. The method of claim 60, wherein the bacterium is an actinomycete and the inhibitor inhibits intracellular production of mycothiol. 67. The method of claim 60, wherein the bacterium is selected from the group consisting of the pathogenic bacteria M. smegmatis, M. tuberculosis, M. leprae, M. bovis, M. intracellulare, M. africanum, M. marinarum, M. chelonai, Corynebacterium diphtheria, Actinomyces israelii, M. avium complex (MAC), M. ulcerans, M abscessus, and M. scrofulaceum. 68. The method of claim 60, wherein the bacterium is an actinomycete. 69. A method for inhibiting growth of an acetyl glucosaminyl inositol-producing bacterium in a mammal, said method comprising administering to the mammal an effective amount of an inhibitor of intracellular acetyl glucosaminyl inositol deacetylase, thereby inhibiting growth of the bacterium in the mammal. 70. The method of claim 69, wherein the inhibitor is derived from GlcN-Ins by replacing the amino group therein with a moiety that chelates a metal ion selected from the group consisting of Mn2+ and Ni2, Cd2+, Co2+, and Zn2+, or otherwise binds the enzyme active site in the deacetylase. 71. The method of claim 70, wherein the moiety is ClCH2CONH—. 72. The method of claim 70 wherein the moiety is HONHCONH—. 73. The method of claim 70, wherein the moiety is HONHCOCH2—. 74. The method of claim 70, wherein the moiety is HOPO(CH3)NH—. 75. The method of claim 70, wherein the moiety is HOPO(CH3)nCH2—; wherein n=1-5. 76. The method of claim 70 wherein the moiety is HSCH2(CH2)nNH—; wherein n=2-5. 77. The method of claim 70 wherein the moiety is HS(CH2)nCONH—; wherein n=1-3. 78. The method of claim 69 wherein the acetyl glucosaminyl inositol-producing bacterium is a mycothiol-producing bacterium. 79. A process for preparation of 1-D-myo-inosityl-2-amino-2-deoxy-α-D-glucopyranoside (GlcN-Ins), said method comprising: contacting an acyl glucosaminyl inositol with an deacetylase of claim 1 under suitable conditions so as to hydrolyze the amide bond therein, and obtaining the GlcN-Ins. 80. A method for determination of acetyl glucosaminyl inositol (GlcNAc-Ins) in a sample, said method comprising: a) contacting a sample containing GlcNAc-Ins with a deacetylase of claim 1 under suitable conditions and b) determining the amount of GlcN-Ins produced, wherein the amount of GlcN-Ins produced is a measure of the GlcNAc-Ins in the sample. 81. The method of claim 80, wherein the amount of GlcN-Ins is determined by HPLC after labeling thereof with a fluorometric or calorimetric reagent. 82. The method of claim 81, wherein the fluorometric reagent is fluorescamine. 83. The method of claim 81, wherein the fluorometric reagent is 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate.

<SOH> BACKGROUND INFORMATION <EOH>Glutathione (GSH) is the dominant low molecular weight thiol in most eukaryotes and Gram-negative bacteria, and it plays a key role in protection of the cell against oxygen toxicity and electrophilic toxins. However, most gram positive bacteria, including many strict aerobes, do not produce glutathione. Yet aerobic organisms are subjected to oxidative stress from many sources, including atmospheric oxygen, basal metabolic activities, and, in the case of pathogenic microorganisms, toxic oxidants from the host phagocytic response intended to destroy the bacterial invader. Actinomycetes, including Streptomyces and Mycobacteria , do not make GSH but produce instead millimolar levels of mycothiol (MSH, AcCys-GlcN-Ins), an unusual conjugate of N-acetylcysteine (AcCys) with 1-D-myo-inosityl-2-amino-2-deoxy-α-D-glucopyranoside (GlcN-Ins). The biochemistry of mycothiol appears to have evolved completely independently of that of glutathione. However, it has already been established that the metabolism of mycothiol parallels that of glutathione metabolism in two enzymatic processes. First, formaldehyde is detoxified in glutathione-producing organisms by NAD/glutathione-dependent formaldehyde dehydrogenase (L. Uotila, et al. (1989) in Glutathione: Chemical, Biochemical, and Medical Aspects—Part A (D. Dolphin, et al., Eds.) pp 517-551, John Wiley & Sons, et al.). An analogous process involving NAD/mycothiol-dependent formaldehyde dehydrogenase has been identified in the actinomycete Amycolatopsis methanolica (M. Misset-Smits, et al. (1997) FEBS Lett. 409:221-222). This enzyme has been sequenced (A. Norin, et al. (1997) Eur. J. Biochem. 248:282-289). The second enzymatic process involves a mycothiol homolog of glutahione reductase recently cloned from M. tuberculosis and expressed in M. smegmatis (M. P. Patel, et al. (1999) J. Amer. Chem. Soc. 120:11538-11539; M. P. Patel et al. (1999) Biochemistry 38:11827-11833; M. P. Patel et al (2001) Biochemistry 40:3119-3126). The reductase is reasonably specific for the disulfide of mycothiol, but is also active with the disulfide of AcCys-GlcN, the desmyo-inositol derivative of mycothiol. Therefore, there is a need in the art for methods and compounds useful for investigation of the details of the metabolism of mycothiol and comparison with the established roles for the metabolism of glutathione. Antibiotic resistance of pathogenic bacteria, including pathogenic actinomycetes, such as M. tuberculosis , is a well-known problem faced by medical practitioners in treatment of bacterial diseases. Therefore, there is a need in the art for new antibiotics, drugs and vaccines and for screening techniques to discover antibiotics, drugs and vaccines effective to treat or prevent bacterial infections in humans and in other mammals, such as domestic and farm animals.

<SOH> SUMMARY OF THE INVENTION <EOH>The present invention solves these and other problems in the art by providing, in one embodiment, purified acetyl glucosaminyl inositol deacetylases, characterized as having an N-terminal region with an amino acid sequence with 40% or more sequence identity to SEQ ID NO:2 and conservative variations thereof, four domains of conservation, wherein three of the domains contain conserved histidine residues, and deacetylase activity against acetyl glucosaminyl inositol. In another embodiment, the invention provides purified acetyl glucosaminyl inositol deacetylases characterized as having an N-terminal region with an amino acid sequence with at least 40% sequence identity to SEQ ID NO:2, one or more domains of conservation containing conserved metal chelating residues, and deacetylase activity against acetyl glucosaminyl inositol in the presence of metal ion. Antibodies and functional fragment thereof, that bind specifically to invention deacetylases, isolated polynucleotide that encode the invention deacetylases, vectors containing a polynucleotide that encodes an invention deacetylase and cells containing such vectors are also provided. In another embodiment, the invention provides methods for identifying an inhibitor of acetyl glucosaminyl inositol deacetylase by contacting a candidate compound with an invention deacetylase in the presence of an acyl glucosaminyl inositol under suitable conditions and determining the presence or absence of breakdown products of the acyl glucosaminyl inositol indicative of deacetylase activity. Substantial absence of the breakdown product of deacetylase activity is indicative of a candidate compound that inhibits activity of the deacetylase. In yet another embodiment, the present invention provides high throughput screening methods for identifying inhibitors of the invention deacetylases. The screening methods involve contacting each of a plurality of candidate compounds with an invention deacetylase in the presence of acyl glucosaminyl inositol under suitable conditions to form a plurality of reaction mixtures and determining the presence or absence of binding to the reaction mixtures of a detectable marker that binds to free amine. Substantial absence of binding of the marker to a reaction mixture in the invention screening methods is indicative of a candidate compound that inhibits activity of the deacetylase. In yet another embodiment, the present invention provides derivatives of 1-D-myo-inosityl-2-amino-2-deoxy-α-D-glucopyranoside (GlcN-Ins), wherein the derivative contains a reactive residue attached to the amino group that promotes oxidative stress and wherein the derivative is selectively toxic to a mycothiol-producing actinomycete by being concentrated in the actinomycete. In still another embodiment, the present invention provides live mutant actinomycetes. The invention mutant actinomycetes have a genome with a disruption in an endogenous acetyl glucosaminyl inositol deacetylase gene, wherein said disruption prevents function of an endogenous acetyl glucosaminyl inositol deacetylase while cell surface proteins and lipids are substantially unaffected, and wherein the disruption results in the mutant actinomycetes exhibiting survival in mammalian white blood cells for an immune response raising period of time. In yet another embodiment, the present invention provides methods for decreasing the virulence of a pathogenic acetyl glucosaminyl inositol deacetylase-producing bacterium in mammalian cells by introducing into the bacterium an inhibitor of acetyl glucosaminyl inositol deacetylase activity. The intracellular presence of the inhibitor in the bacterium decreases activity of the deacetylase, thereby decreasing mycothiol biosynthesis by the bacterium as compared with untreated control bacterium. By virulence is meant the relative power and degree of pathogenicity possessed by organisms to produce disease as measured by clinical symptoms particular to the disease under consideration. For example, the virulence of a M. tuberculosis is measured with reference to the manifestation in an infected individual of the clinical symptoms recognized by a medical practitioner as indicative of tuberculosis. In another embodiment, the present invention provides inhibitors of the invention acetyl glucosaminyl inositol deacetylases wherein the inhibitor is derived from GlcN-Ins by replacing the amino group therein with a moiety that chelates Zn 2+ , or otherwise binds the enzyme active site of the deacetylase. In still another embodiment, the present invention provides methods for inhibiting growth of an acetyl glucosaminyl inositol-producing bacterium in a mammal, by administering to the mammal an effective amount of an inhibitor of intracellular acetyl glucosaminyl inositol deacetylase, thereby inhibiting growth of the bacterium in the mammal. In yet another embodiment, the present invention provides processes for preparation of 1-D-myo-inosityl-2-amino-2-deoxy-α-D-glucopyranoside (GlcN-Ins) by contacting an acyl glucosaminyl inositol with an invention deacetylase under suitable conditions so as to hydrolyze the amide bond therein, and obtaining the GlcN-Ins.

Process for the electromagnetic modelling of electronic components and systems

Process for the electromagnetic modelling of electronic components and systems, for the extraction of certain electrical parameters, such as the static V-I characteristics and the input and output impedance, the output switching times in particular conditions of load and the transition times of the protection diodes. A test machine of the commercial type is used for these measurements, by generating stimulus signals and measuring the correlated signals, the test machine being suitable for parametric direct current measurements, functional tests and digital integrated circuit timing and also being used as a time domain reflectometer. The measurement phase is followed by a simulation phase during which the electric parameters used for modelling electronic components and systems are extracted.

1. Process for the electromagnetic modelling of electronic components and systems, in particular for the extraction of certain electrical parameters, such as the static V-I characteristics and the input and output impedance, the output switching times in particular conditions of load and the transition times of the protection diodes of said electronic components and systems, to ports of these components and systems suitable stimulus signals are sent and the correlated signals are measured, reconstructing the respective time patterns, characterised in that a test machine of the commercial type is used for generating the stimulus signals and measuring the correlated signals, the test machine being suitable for parametric direct current measurements, functional tests and digital integrated circuit timing and also being used as a time domain reflectometer, in which case the test machine sends said stimulus signals to the component or system inputs or outputs, fitted on a suitable support and connected by means of suitable interconnection channels, and detects the reflection due to impedance offset, providing the respective time patterns, which are then compared with those provided by a simulated measurement system model, comprising the test machine signal generation and measurement part, the interconnection channels and the support, to verify the coincidence between the measure time pattern and that provided by the simulated model obtained by setting different values with respect to the electric parameters to be determined. 2. Process according to claim 1, characterised in that the model simulated by the measurement system is obtained by the reflectometer response of each channel of the test machine in the situation in which the support of the component or system is not connected to the machine. 3. Process according to claim 1, characterised in that the reflected signals are reconstructed by means of the periodical sampling method, by fine tuning the sampling pulse phase and operating on the comparison threshold of the measurement part of the test machine to identify the succession of values assumed in time by the reflected signal, so as to obtain the time pattern. 4. Process according to claim 1, characterised in that for measuring the electronic component and system input and output impedance, a variable load impedance is introduced in the simulated model, so as to obtain various patterns of the reflected signal, following the same stimulus signal, to find the pattern which best reproduces the measured pattern by subsequent approximation, the simulated impedance value corresponding to the impedance of the input or output to be determined. 5. Process according to claim 1, characterised in that for measuring the switching time of the outputs or transition time of the protection diode, a variable time is introduced in the simulated model, so as to obtain different patterns of the reflected signal, given the same stimulus signal, to find the pattern which best reproduces the measured pattern by subsequent approximation, the simulated switching or transition time corresponding to the value to be determined.

Electrophoretic analysis method

By capillary electrophoresis, an effective mobility to play an important role for identifying an unknown sample is quickly determined highly precisely. A standard substance is added to a sample containing an unknown substance (unknown sample)(S101). This unknown sample to which the standard substance is added is introduced into a capillary, and capillary electrophoresis is executed (s103). Next, by executing this capillary electrophoresis, the various parameters of the standard substance and the unknown sample are determined (S107). Next, on the basis of the various parameters and a theoretical equation, the effective mobility of the unknown sample is calculated. On the basis of the calculated effective mobility of the unknown sample, the substance of the unknown sample is identified (S109).

1. An electrophoretic analysis method including the steps of: adding a standard substance to a sample containing an unknown sample and measuring migration times of the standard substance and the unknown sample by electrophoresis and a migration time of a substance having a mobility of 0 or a migration time of a system peak; and determining an effective mobility of the unknown sample on the basis of the migration times of the standard substance and the unknown sample and the migration time of the substance having a mobility of 0 or the migration time of a system peak as measured in the measuring step, and a known effective mobility of the standard substance at a standard temperature. 2. The electrophoretic analysis method according to claim 1, wherein the step of determining an effective mobility of the unknown sample is to determine an effective mobility m0 of the unknown sample on the basis of a migration time tS of the standard substance, a migration time t of the unknown sample, a migration time teof of the substance having a mobility of 0 or of the system peak, and an effective mobility m0, S of the standard substance at the standard temperature according to the following equation: m0={(teof/t−1)/(teof/ts−1)}·mo, s 3. The electrophoretic analysis method according to claim 1, wherein the step of determining an effective mobility of the unknown sample is to determine the effective mobility of the unknown sample on the basis of the migration times of the two or more standard substances, the migration time of the unknown sample, the migration time of the substance having a mobility of 0 or of the system peak, and the effective mobilities of the standard substances at the standard temperature. 4. The electrophoretic analysis method according to claim 3, wherein the step of determining an effective mobility of the unknown sample is to determine an effective mobility m0 of the unknown sample on the basis of migration times tA and tB of arbitrary two substances of the two or more standard substances, a migration time t of the unknown sample, a migration time teof of the substance having a mobility of 0 or of the system peak, and effective mobilities m0, A and m0, B of the standard substances at the standard temperature according to the following equations: m0={(tA−τ)(teof−t)/(t−τ)(teof−tA)}·m0, A τ={m0, AtA(teof−tB)−m0, BtB(teof−tA)}/{m0, A(teof−tB)−m0, B(teof−tA)} 5. The electrophoretic analysis method according claim 1, further including a step of identifying the substance of the unknown sample from the effective mobility of the unknown sample. 6. An electrophoretic analysis method including the steps of: adding a standard substance to a sample containing an unknown sample and measuring migration times of the standard substance and the unknown sample by electrophoresis; and determining an effective mobility of the unknown sample on the basis of the migration times of the standard substance and the unknown sample as measured in the measuring step, and a known effective mobility of the standard substance at a standard temperature. 7. The electrophoretic analysis method according to claim 6, wherein the step of determining an effective mobility of the unknown sample is to determine an effective mobility m0 of the unknown sample on the basis of migration times tA, tB and tC of arbitrary three substances of the three or more standard substances, a migration time t of the unknown sample, and effective mobilities m0, A and m0, B and m0, C of the standard substances at the standard temperature according to the following equations: m0={(ti−t)(tj−τ)(m0, j−m0, i)}/{(t−τ)(ti−tj)}+m0, i wherein i and j are any one of the standard substances A, B and C, and i is not equal to j, τ={m0, AtA(tC−tB)+m0, BtB(tA−tC)+m0, CtC(tB−tA)}/{m0, A(tC−tB)+m0, B(tA−tC)+m0, C(tB−tA)} 8. The electrophoretic analysis method according to claim 2 further including a step of identifying the substance of the unknown sample from the effective mobility of the unknown sample. 9. The electrophoretic analysis method according to claim 3 further including a step of identifying the substance of the unknown sample from the effective mobility of the unknown sample. 10. The electrophoretic analysis method according to claim 4 further including a step of identifying the substance of the unknown sample from the effective mobility of the unknown sample.

<SOH> BACKGROUND OF THE INVENTION <EOH>In recent years, in the case where polymers, colloid particles, etc. contained in various solutions are identified, an analysis method utilizing a phenomenon in which the polymers or colloid particles move (migrate) corresponding to potential differences, so-called electrophoresis, become widespread. The migration velocity of the polymers or colloid particles in the solution that is influenced by not only the kind and concentration of an electrolyte in the solution but also the shape and size of the particles of the polymers or colloid particles themselves. For this reason, capillary zone electrophoresis (CZE) that is one of the analysis methods utilizing this electrophoresis is being watched as, for example, an important analysis method for analyzing the length of DNA fragments contained in proteins, etc. This capillary zone electrophoretic analysis method is required to have high reproducibility regarding a peak position to be used directly as a quantitative index of an unknown substance (unknown sample) in a solution. In the case where the analysis results of this capillary zone electrophoresis is expressed by a pherogram based on coordinate axes in which an abscissa represents the migration time by an unknown sample in a solution, and an ordinate represents the peak position, in order to obtain a peak position with good reproducibility, it is a very important factor to control each of a migration voltage (current), a temperature of a separation chamber, and electroosmotic flow. It is easily possible to control the migration voltage (current) and the temperature of the separation chamber by making the analysis conditions (such as an inner diameter of a capillary and a supporting electrolyte) identical. On the other hand, a temperature increase of a separation tube by a Joule's heat is inevitable and the control of the electroosmotic flow becomes difficult. For these reasons, the reproducibility of the electrophoresis of a usual capillary zone electrophoretic pherogram having a time axis is not high as compared with that in high-performance liquid chromatography (HPLC). As factors of disturbing the reproducibility of the migration time of an unknown sample in a solution are enumerated a change of electroosmotic flow, a relaxation effect of potential gradient (RPG), and a temperature increase by a Joule's heat generated with the application of a high voltage. Since these occur due to complex factors, it is difficult to control them. In the case where the measurement is carried out by using different equipments, it is assumed that even if the sample or supporting electrolyte is identical, it is difficult to obtain the same migration time. Accordingly, in order to correctly compare the measurement results in the case of using different equipments, it is necessary to standardize the electrophoresis data. On the other hand, Lee, et al. proposed a migration index (MI) using a value obtained by dividing a quantity of electricity, i.e., an integral value of current, by a cross-sectional area of a capillary and a whole length of the capillary and an adjusted migration index (AMI) obtained by correcting MI of the osmotic flow (the units of MI and AMI in the description being “μC/m 3 ”). It was considered that if the supporting electrolyte is identical, AMI could become a considerably good quantitative index. However, since a conductivity of the supporting electrolyte is included as one of the parameters of AMI, even a slight difference in the composition (for example, the case where even when a pH is substantially the same, counter ions are different) produces different measurement results.

<SOH> BRIEF SUMMARY OF THE INVENTION <EOH>In order to prevent the matters as described above, it may be required to propose an electrophoretic analysis method using an effective mobility relying upon only conditions of a sample ion and a supporting electrolyte as a quantitative index. However, in the conventional conversion from the migration time to the effective mobility, in many cases, the change of the mobility by a Joule's heat and the relaxation effect of potential gradient were not taken into consideration. Here, the technology related to the invention will be described. As the method of correcting the change of the mobility by a Joule's heat and the relaxation effect of potential gradient, the present inventors previously filed an application for patent with respect to a method in which a standard substance is added to an unknown sample, capillary zone electrophoresis is executed, a migration time of the standard substance is determined, and assuming that an osmotic flow velocity linearly changes with respect to the time, the migration time of the standard substance is converted into an effective mobility (Japanese Patent Application No. 11-251604). This method does not correct temperature dependency of the effective mobility of the sample (temperature dependency of a sample ion itself and temperature dependency to a supporting electrolyte) and so on in relation with a strict physical phenomenon but uses a hypothetic osmotic flow velocity on the assumption that the osmotic flow velocity linearly changes with respect to the time. Accordingly, it is assumed that all of phenomena are included in coefficients in empirical equations representing this linear change, whereby physical meanings become vague. Thus, the present inventors grasped three factors that lower the reproducibility (i.e., temperature dependency of the sample ion itself influencing the migration velocity, temperature dependency to the supporting electrolyte, and a delay of the migration time based on the relaxation effect of potential gradient) as original physical phenomena and groped for a method of theoretically eliminating these factors. Under these circumstances, the invention is aimed to quickly determine with high precision an effective mobility to play an important role for identifying an unknown sample by capillary zone electrophoresis. Further, the invention is aimed to eliminate a scattering of the effective mobility by operational conditions of an analysis equipment and to enhance reliability of data as a standardized index of every unknown sample. Moreover, the invention is aimed to use the same index in different analysis equipments. According to the first dissolution means of the invention, there is provided an electrophoretic analysis method including: a step of adding a standard substance to a sample containing an unknown sample and measuring migration times of the standard substance and the unknown sample by electrophoresis and a migration time of a substance having a mobility of 0 or a migration time of a system peak, and a step of determining an effective mobility of the unknown sample on the basis of the migration times of the standard substance and the unknown sample and the migration time of the substance having a mobility of 0 or the migration time of a system peak as measured in the foregoing measuring step, and a known effective mobility of the standard substance at a standard temperature. According the second dissolution means of the invention, there is provided an electrophoretic analysis method including: a step of adding a standard substance to a sample containing an unknown sample and measuring migration times of the standard substance and the unknown sample by electrophoresis, and a step of determining an effective mobility of the unknown sample on the basis of the migration times of the standard substance and the unknown sample as measured in the foregoing measuring step, and a known effective mobility of the standard substance at a standard temperature. A more detailed explanation of the invention is provided in the following description and appended claims take in conjunction with the accompanying drawings.

Single molecule sequencing method

The invention relates to a method for single molecule sequencing of nucleic acids and to a device suitable for carrying out said method.

1-36. (canceled) 37. A method for sequencing nucleic acids, comprising: (a) providing a support particle on which a nucleic acid molecule has been immobilized, with essentially all nucleotide building blocks of at least one base type in at least one strand of said nucleic acid molecule carrying a fluorescent label, (b) introducing said support particle into a sequencing device comprising a micro channel, (c) arresting said support particle in said sequencing device, (d) progressively removing by cleavage individual nucleotide building blocks from the immobilized nucleic acid molecule, (e) passing the removed nucleotide building blocks through a microchannel by means of a hydrodynamic flow and (f) determining the base sequence of said nucleic acid molecule based on the sequence of said removed nucleotide building blocks. 38. The method as claimed in claim 37, wherein said support particle is made of a material selected form the group consisting of plastic, glass, quartz, metal, semimetal, metal oxides and a composite material. 39. The method as claimed in claim 37, wherein the diameter of the support particle is from 0.5 to 10 μm. 40. The method as claimed in claim 37, wherein the nucleic acid molecule is immobilized on the support particle via it 5′-terminus by means of bioaffinity interactions. 41. The method as claimed in claim 40, wherein a 5′-biotinylated nucleic acid molecule is immobilized to an avidine- or streptavidine-coated support particle. 42. The method as claimed in claim 37, wherein the nucleic acid molecule is immobilized in single-stranded form on the support particle. 43. The method as claimed in claim 37, wherein the nucleic acid molecule molecule is immobilized in double-stranded form on the support particle, it being possible for labeled nucleotide building blocks to be removed by cleavage only from one single strand. 44. The method as claimed in claim 37, wherein essentially all nucleotide building blocks of at least two base types carry a fluorescent label. 45. The method as claimed in claim 37, wherein the support particle is arrested using a capturing laser. 46. The method as claimed in claim 37, wherein the support particles are arrested in a microchannel. 47. The method as claimed in claim 37, wherein individual nucleotide building blocks are removed by cleavage by an exonuclease. 48. The method as claimed in claim 47, wherein T7 DNA polymerase, E. coli exonuclease I or E. coli exonuclease III is used. 49. The method as claimed in claim 37, wherein the removed nucleotide building blocks are passed through a microchannel having a diameter of from 1 to 100 μm. 50. The method as claimed in claim 37, wherein the removed nucleotide building blocks are passed through a microchannel with a velocity of from 1 to 50 mm/s. 51. The method as claimed in claim 37, wherein the determination is carried out by means of confocal fluorescence measurement in a detection volume element. 52. The method as claimed in claim 51, wherein the determination is carried out by means of confocal single molecule detection, such as, for example, fluorescence correlation spectroscopy. 53. The method as claimed in claim 37, wherein the determination is carried out by means of a time-resolved decay measurement or time gating in a detection volume element. 54. The method as claimed in claim 37, wherein nucleic acid molecules are determined in parallel in a plurality of microchannels. 55. The method as claimed in claim 37, wherein a sequence with known base sequence is attached to the nucleic acid molecule to be sequenced. 56. The method as claimed in claim 37, wherein the support particles are arrested essentially in the center of the microchannel and the removed nucleotide building blocks are directed in a laminar flow to a detection volume element which has been positioned in the center of the channel. 57. The method as claimed in claim 56, wherein the detection volume element is kept as small as possible in order to only just detect all removed nucleotide building blocks. 58. A device for sequencing an analyte in a sample fluid, which comprises: (a) an optically transparent microchannel, (b) means for introducing a support particle on which a nucleic acid molecule has been immobilized into said microchannel, with essentially all nucleotide building blocks of at least one base type in at least one strand of said nucleic acid molecule carrying a fluorescent label, (c) means for arresting said support particle at a predetermined position in said microchannel, (d) means for generating a hydrodynamic flow in said microchannel, (e) means for progressively removing cleavage individual nucleotide building blocks from the immobilized nucleic acid molecule, and (f) means for sequentially detecting the removed nucleotide building blocks. 59. A method for sequencing nucleic acids, which comprises the following steps: (a) providing a support particle on which a nucleic acid molecule ahs been immobilized, with essentially all nucleotide building blocks in at least one strand of said nucleic acid molecule carrying a fluorescent label and with 2 fluorescent labels with different properties being used for the 4 bases. (b) introducing said support particle into a sequencing device comprising a microchannel, (c) arresting said support particle in said sequencing device. (d) progressively removing by cleavage individual nucleotide building blocks from the immobilized nucleic acid molecule, (e) passing the removed nucleotide building blocks through a microchanel and (f) determining the base sequence of said nucleic acid molecule based on the sequence of said removed nucleotide building blocks. 60. The method as claimed in claim 59, wherein the different spectroscopic properties are selected from emission wavelength or/and lifetime. 61. The method as claimed in claim 59, wherein step (a) comprises providing at least 2 support particles on which nucleic acid molecules have been immobilized which have at least partially overlapping sequences and in which the 2 fluorescent labels, in each case, have been assigned to different bases or/and base combinations. 62. The method as claimed in claim 61, wherein a first support particle in each case a first fluorescent label is assigned to 2 bases, B1 and B2, and a second fluorescent label is assigned to B3 and B4, on a second support particle in each case a first fluorescent labile is assigned to 2 bases, B1 and B3, and a second fluorescent label is assigned to 2 bases, B2 and B4, and, where appropriate, on a third support particle in each case a first fluorescent label is assigned to 2 bases, B1 and B4, and a second fluorescent label is assigned to 2 bases, B2 and B3. 63. The method as claimed in claim 61, wherein on a first support particle in each case a first fluorescent label is assigned to a base B1 and a second fluorescent label is assigned to 3 bases, B2, B3 and B4, on a second support particle in each case a first fluorescent label is assigned to a base B2 and a second fluorescent label is assigned to 3 bases, B1, B3 and B4, on a third support particle in each case a first fluorescent label is assigned to a base B3 and a second fluorescent label is assigned to 3 bases, B1, B2 and B4, and on a fourth support particle in each case a first fluorescent label is assigned to a base B4 and a second fluorescent label is assigned to 3 bases, B1, B2 and B3. 64. The method as claimed in claim 61, wherein a first support particle a first fluorescent label is assigned to 2 bases and a second fluorescent label is assigned to the other 2 bases, and on a second support particle a first fluorescent label is assigned to one base and a second fluorescent label is assigned to the other three bases. 65. A method for sorting particles in a microchannel, which comprises the following steps: (a) passing particles through a detection element in said microchannel, said detection element being adjusted in such a way that a capturing laser is activated if a predetermined parameter is present on the particle and that said capturing laser is not activated if said predetermined parameter is not present on said particle, (b) arresting a particle on which said predetermined parameter is present by said capturing laser in a measuring element, (c) interrupting the sorting process and (d) measuring the arrested particle. 66. A device for sequencing nucleic acids, comprising: (a) an optically transparent microchannel, (b) means for introducing a support particle on which a nucleic acid molecule has been immobilized into said microchannel, with essentially all nucleotide building blocks of at least one base type in at least one strand of said nucleic acid molecule carrying a fluorescent label, (c) means for arresting said support particle at a first predetermined position in said microchannel, (d) means for transporting said support particle to a second predetermined position of said mircochannel, (e) means for generating a flow in said microchannel, (f) means for progressively removing by cleavage individual nucleotide building blocks from the immobilized nucleic acid molecule at the second position, and (g) means for sequentially detecting the removed nucleotide building blocks. 67. A device for sequencing nucleic acids, comprising: (a) a support comprising a system of microchannels which are in fluid communication with one another, (b) an opening for introducing a support particle having a nucleic acid molecule immobilized thereon into a microchannel, with essentially all nucleotide building blocks of at least one base type in at least one strand of said nucleic acid molecule carrying a fluorescent label, (c) an opening for feeding a nucleic acid-degrading enzyme into a microchannel, (d) a plurality of openings for discharging fluid from said support, (e) an opening for feeding buffer into a microchannel, (f) means for generating a liquid stream in said microchannels, (g) means for capturing the support particle at a first predetermined position, (h) means for transporting a captured support particle to a second predetermined position, (i) means for progressively removing by cleavage individual nucleotide building blocks from the immobilized nucleic acid molecule at the second predetermined position, and (j) means for sequentially detecting the removed nucleotide building blocks. 68. The device as claimed in claim 67, wherein the diameter of the microchannels leading to the discharge openings is larger, preferably at least 1.5 times larger, than the diameter of the microchannels leading to the feeding openings and: 69. The device as claimed in claim 66, wherein the means (f) for generating a hydrodynamic flow are provided in the support. 70. The device as claimed in claim 66, wherein means for applying an electric field between the second predetermined position and the first predetermined position are provided. 71. A method for sequencing nucleic acids, characterized in that a device as claimed in claim 66, is used. 72. A method for sequencing nucleic acids, comprising: (a) providing a support particle on which a nucleic acid molecule has been immobilized, with essentially all nucleotide building blocks of at least one base type in at least one strand of said nucleic acid molecule carrying a fluorescent label, (b) introducing said support particle into an opening of a support comprising a system of microchannels which are in fluid communication with one another, (c) arresting said support particle at a first predetermined position within said support, (d) transporting said support particle to a second predetermined position within said support, (e) progressively removing by cleavage individual nucleotide building blocks from the immobilized nucleic acid molecule at said second predetermined position, (f) passing the removed nucleotide building blocks through a microchannel, and (g) determining the base sequence of said nucleic acid molecule based on the sequence of said removed nucleotide building blocks. 73. The method as claimed in claim 38, wherein the diameter of the support particle is from 0.5 to 10 μm. 74. The method as claimed in claim 60, wherein step (a) comprises providing at least 2 support particles on which nucleic acid molecules have been immobilized which have at least partially overlapping sequences and in which the 2 fluorescent labels, in each case, have been assigned to different bases or/and base combinations.

Methods and materials relating to carcinoembryonic antigen-like (cea-like) polypeptides and polynucleotides

The invention provides novel polynucleotides and polypeptides encoded by such polynucleoddes and mutants or variants thereof that correspond to a novel human secreted CEA-like polypeptide. These polynucleotides comprise nucleic acid sequences isolated from cDNA library from rectum (Hyseq clone identification numbers 15456780 (SEQ ID NO: 1)). Other aspects of the invention include vectors containing processes for producing novel human secreted CEA-like polypeptides, and antibodies specific for such polypeptides.

1. An isolated polynucleotide comprising a nucleotide sequence selected from the group consisting of SEQ ID NO: 2-3 and 5, the translated protein coding portion thereof, the mature protein coding portion thereof, the extracellular portion thereof, or the active domain thereof. 2. An isolated polynucleotide encoding a polypeptide with biological activity, which polynucleotide hybridizes to the complement of a polynucleotide of claim 1 under stringent hybridization conditions. 3. An isolated polynucleotide encoding a polypeptide with biological activity, said polynucleotide having greater than about 90% sequence identity with the polynucleotide of claim 1. 4. The polynucleotide of claim 1 which is a DNA sequence. 5. An isolated polynucleotide which comprises the complement of the polynucleotide of claim 1. 6. A vector comprising the polynucleotide of claim 1. 7. An expression vector comprising the polynucleotide of claim 1. 8. A host cell genetically engineered to express the polynucleotide of claim 1. 9. The host cell of claim 8 wherein the polynucleotide is in operative association with a regulatory sequence that controls expression of the polynucleotide in the host cell. 10. An isolated polypeptide comprising an amino acid sequence which is at least 80% identical to the amino acid sequence selected from the group consisting of SEQ ID NO: 4 and 6-10, the translated protein coding portion thereof, the mature protein coding portion thereof, the extracellular portion thereof, or the active domain thereof. 11. A composition comprising the polypeptide of claim 10 and a carrier. 12. A polypeptide, having CEA-like activity, comprising at least ten consecutive amino acids from the polypeptide sequences selected from the group consisting of SEQ ID NO: 4 and 6-10. 13. The polypeptide of claim 12, comprising at least five consecutive amino acids from the polypeptide sequences selected from the group consisting of SEQ ID NO. 4 and 6-10. 14. A polynucleotide encoding a polypeptide according to claim 12. 15. A polynucleotide encoding a polypeptide according to claim 13. 16. A polynucleotide encoding a polypeptide according to claim 10. 17. An antibody specific for the polypeptide of claim 10. 18. A method for detecting the polynucleotide of claim 1 in a sample, comprising: a) contacting the sample with a compound that binds to and forms a complex with the polynucleotide of claim 1 for a period sufficient to form the complex; and b) detecting the complex, so that if a complex is detected, the polynucleotide of claim 1 is detected. 19. A method for detecting the polynucleotide of claim 1 in a sample, comprising: a) contacting the sample under stringent hybridization conditions with nucleic acid primers that anneal to the polynucleotide of claim 1 under such conditions; b) amplifying a product comprising at least a portion of the polynucleotide of claim 1; and c) detecting said product and thereby the polynucleotide of claim 1 in the sample. 20. The method of claim 19, wherein the polynucleotide comprises an RNA molecule and the method further comprises reverse transcribing an annealed RNA molecule into a cDNA polynucleotide. 21. A method for detecting the polypeptide of claim 10 in a sample, comprising: a) contacting the sample with a compound that binds to and forms a complex with the polypeptide under conditions and for a period sufficient to form the complex; and b) detecting formation of the complex, so that if a complex formation is detected, the polypeptide of claim 10 is detected. 22. A method for identifying a compound that binds to the polypeptide of claim 10, comprising: a) contacting the compound with the polypeptide of claim 10 under conditions and for a time sufficient to form a polypeptide/compound complex; and b) detecting the complex, so that if the polypeptide/compound complex is detected, a compound that binds to the polypeptide of claim 10 is identified. 23. A method for identifying a compound that binds to the polypeptide of claim 10, comprising: a) contacting the compound with the polypeptide of claim 10, in a cell, for a time sufficient to form a polypeptide/compound complex, wherein the complex drives expression of a reporter gene sequence in the cell; and b) detecting the complex by detecting reporter gene sequence expression, so that if the polypeptide/compound complex is detected, a compound that binds to the polypeptide of claim 10 is identified. 24. A method of producing a CEA-like polypeptide, comprising, a) culturing the host cell of claim 8 under conditions sufficient to express the polypeptide in said cell; and b) isolating the polypeptide from the cell culture or cells of step (a). 25. A kit comprising the polypeptide of claim 10. 26. A nucleic acid array comprising the polynucleotide of claim 1 or a unique segment of the polynucleotide of claim 1 attached to a surface. 27. The array of claim 26, wherein the array detects full-matches to the polynucleotide or a unique segment of the polynucleotide of claim 1. 28. The array of claim 26, wherein the array detects mismatches to the polynucleotide or a unique segment of the polynucleotide of claim 1. 29. A method of treatment of a subject in need of enhanced activity or expression of CEA-like polypeptide of claim 10 comprising administering to the subject a composition selected from the group consisting of: (a) a therapeutic amount of a agonist of said polypeptide; (b) a therapeutic amount of the polypeptide; and (c) a therapeutic amount of a polynucleotide encoding the polypeptide in a form and under conditions such that the polypeptide is produced, and a pharmaceutically acceptable carrier. 30. A method of treatment of a subject having need to inhibit activity or expression of CEA-like polypeptide of claim 10 comprising administering to the subject a composition selected from the group consisting of: (a) a therapeutic amount of an antagonist to said polypeptide; (b) a therapeutic amount of a polynucleotide that inhibits the expression of the nucleotide sequence encoding said polypeptide; and (c) a therapeutic amount of a polypeptide that competes with the CEA-like polypeptide for its ligand and a pharmaceutically acceptable carrier.

<SOH> 2. BACKGROUND ART <EOH>Identified polynucleotide and polypeptide sequences have numerous applications in, for example, diagnostics, forensics, gene mapping; identification of mutations responsible for genetic disorders or other traits, to assess biodiversity, and to produce many other types of data and products dependent on DNA and amino acid sequences. Proteins are known to have biological activity, for example, by virtue of their secreted nature in the case of leader sequence cloning, by virtue of their cell or tissue source in the case of PCR-based techniques, or by virtue of structural similarity to other genes of known biological activity. It is to these polypeptides and the polynucleotides encoding them that the present invention is directed. In particular, this invention is directed to novel soluble CEA-like polypeptides and polynucleotides. Many tumors express genes whose products are required for either inducing or maintaining the malignant state (Abbas et al (2000) Cellular and Molecular Immunology, Saunders (Publishers) pp 386). These proteins may serve as markers for the tumor detection or even may provide therapeutic targets. Carcinoembryonic antigens (CEAs, e.g. CD66a-CD66d) were first described as antigens expressed on many carcinomas of colon, pancreas, stomach, and breast. However, it has become apparent with more sensitive detection techniques that these proteins are also expressed during inflammation and also, in minute quantities in normal tissues. Carcinoembryonic antigens are integral membrane glycoproteins belonging to immunoglobulin (Ig) superfamily of receptors. CEA cell adhesion molecule (CEA-CAM), also known as billiary glycoprotein (BGP) and CD66a, is a protein of about 85 kDa and, is highly glycosylated and exhibits at least two tissue specific, alternatively spliced, variants. Immunoglobulin superfamily members that serve as receptors can be classified into three groups according to their cytoplasmic domain characteristics. Transmembrane molecules with immunoreceptor tyrosine activation motifs (ITAMs) (YxxL where x is any amino acid) are usually activating receptors. Those possessing immunoreceptor tyrosine inhibition motifs (ITIMs) (I/L/VxxYxxL/V where x is any amino acid) are inhibitory in nature. There appears to be a third class of short transmembrane receptors like LIR-4, or alternately spliced soluble forms of FDF03 that have no known activating or inhibitory motifs. These molecules by their virtue of extracellular MHC binding domain could function as a “molecular sink” and could inhibit the functions of cognate transmembrane receptors. Several functions have been attributed to CEA. The first Ig domain of CD66a serves as an adhesive module to bind E-selectin and initiate the inflammatory cascade. The mouse CEAs have been shown to be the receptors for mouse hepatitis virus, whereas human CEA has been shown to be a receptor for bacterial proteins from Neisseria gonorrhoeae, Salmonella , and Escherichia coli . More recently, CEA has been shown to be a negative regulator, and therefore a tumor suppressor protein for colonic, prostate and breast carcinomas (Huber et al (1999) J. Biol. Chem. 274, 335-344). CEA-CAM or CD66a has been implicated in transducing signals by its cytoplasmic domain. Several physiological events promote the phosphorylation of tyrosines in the cytoplasmic domain of CEA. It is also reported that stimulation of BGP1 in neutrophils leads to activation of Rac1, PAK, and Jun N-terminal kinase. Similarly, it is reported that the ITIM sequences in the BGP1 cytoplasmic domains interact with protein-tyrosine phosphatases SHP-1 and SHP-2 in epithelial cells (Huber et al (1999) J. Biol. Chem. 274, 335-344). CEA expression is greatly increased in colonic, pancreatic, gastric, and breast carcinomas resulting in a rise in serum levels. Furthermore, post-translational processing of CEA may be altered in tumor cells. Serum CEA is accordingly used to monitor the occurrence or recurrence of metastatic carcinoma after primary treatment. CEA functions as an intercellular adhesion molecule, promoting the binding of tumor cells to one another. Thus CEA may play a role in the way tumor cells interact with one another and with tissues in which they are growing. Thus, CEA appears to be involved in cell adhesion and subsequent signal transduction during normal fetal development and also during inflammation and carcinogenesis. Polynucleotides encoding CEA and polypeptides thereof could serve as potential therapeutics in the treatment of breast, prostate, colon and other cancers. CEA and compounds which bind to CEA could also be useful in treating disorders relating to inflammation and autoimmunity. Soluble CEA could also be used as immunosuppressant in organ transplant patients. Soluble CEA molecule could serve as a decoy receptor in above mentioned bacterial and viral infections.

<SOH> 3. SUMMARY OF THE INVENTION <EOH>This invention is based on the discovery of novel CEA-like polypeptides, novel isolated polynucleotides encoding such polypeptides, including recombinant DNA molecules, cloned genes or degenerate variants thereof, especially naturally occurring variants such as allelic variants, antisense polynucleotide molecules, and antibodies that specifically recognize one or more epitopes present on such polypeptides, as well as hybridomas producing such antibodies. Specifically, the polynucleotides of the present invention are based on a CEA-like polynucleotide isolated from a cDNA library prepared from rectum (Hyseq clone identification numbers 15456780 (SEQ ID NO: 1). The compositions of the present invention additionally include vectors such as expression vectors containing the polynucleotides of the invention, cells genetically engineered to contain such polynucleotides and cells genetically engineered to express such polynucleotides. The compositions of the invention provide isolated polynucleotides that include, but are not limited to, a polynucleotide comprising the nucleotide sequence set forth in the SEQ ID NO: 1-3 or 5; or a fragment of SEQ ID NO: 1-3 or 5; a polynucleotide comprising the full length protein coding sequence of the SEQ ID NO: 1-3 or 5 (for example, SEQ ID NO: 4); and a polynucleotide comprising the nucleotide sequence of the mature protein coding sequence of any of SEQ ID NO: 1-3 or 5. The polynucleotides of the present invention also include, but are not limited to, a polynucleotide that hybridizes under stringent hybridization conditions to (a) the complement of any of the nucleotide sequences set forth in SEQ ID NO: 1-3 or 5; (b) a nucleotide sequence encoding any of SEQ ID NO: 4, 6-10; a polynucleotide which is an allelic variant of any polynucleotides recited above having at least 70% polynucleotide sequence identity to the polynucleotides; a polynucleotide which encodes a species homolog (e.g. orthologs) of any of the peptides recited above; or a polynucleotide that encodes a polypeptide comprising a specific domain or truncation of the polypeptide comprising SEQ ID NO: 4. A collection as used in this application can be a collection of only one polynucleotide. The collection of sequence information or unique identifying information of each sequence can be provided on a nucleic acid array. In one embodiment, segments of sequence information are provided on a nucleic acid array to detect the polynucleotide that contains the segment. The array can be designed to detect full-match or mismatch to the polynucleotide that contains the segment. The collection can also be provided in a computer-readable format. This invention further provides cloning or expression vectors comprising at least a fragment of the polynucleotides set forth above and host cells or organisms transformed with these expression vectors. Useful vectors include plasmids, cosmids, lambda phage derivatives, phagemids, and the like, that are well known in the art. Accordingly, the invention also provides a vector including a polynucleotide of the invention and a host cell containing the polynucleotide. In general, the vector contains an origin of replication functional in at least one organism, convenient restriction endonuclease sites, and a selectable marker for the host cell. Vectors according to the invention include expression vectors, replication vectors, probe generation vectors, and sequencing vectors. A host cell according to the invention can be a prokaryotic or eukaryotic cell and can be a unicellular organism or part of a multicellular organism. The compositions of the present invention include polypeptides comprising, but not limited to, an isolated polypeptide selected from the group comprising the amino acid sequence of SEQ ID NO: 4, 6-10; or the corresponding full length or mature protein. Polypeptides of the invention also include polypeptides with biological activity that are encoded by (a) any of the polynucleotides having a nucleotide sequence set forth in the SEQ ID NO: 1-3 or 5; or (b) polynucleotides that hybridize to the complement of the polynucleotides of (a) under stringent hybridization conditions. Biologically or immunologically active variants of any of the protein sequences listed as SEQ ID NO: 4, 6-10 and substantial equivalents thereofthat retain biological or immunological activity are also contemplated. The polypeptides of the invention may be wholly or partially chemically synthesized but are preferably produced by recombinant means using the genetically engineered cells (e.g. host cells) of the invention. The invention also provides compositions comprising a polypeptide of the invention. Pharmaceutical compositions of the invention may comprise a polypeptide of the invention and an acceptable carrier, such as a hydrophilic, e.g., pharmaceutically acceptable, carrier. The invention also relates to methods for producing a polypeptide of the invention comprising culturing host cells comprising an expression vector containing at least a fragment of a polynucleotide encoding the polypeptide of the invention in a suitable culture medium under conditions permitting expression of the desired polypeptide, and purifying the protein or peptide from the culture or from the host cells. Preferred embodiments include those in which the protein produced by such a process is a mature form of the protein. Polynucleotides according to the invention have numerous applications in a variety of techniques known to those skilled in the art of molecular biology. These techniques include use as hybridization probes, use as oligomers, or primers, for PCR, use in an array, use in computer-readable media, use for chromosome and gene mapping, use in the recombinant production of protein, and use in generation of antisense DNA or RNA, their chemical analogs and the like. For example, when the expression of an mRNA is largely restricted to a particular cell or tissue type, polynucleotides of the invention can be used as hybridization probes to detect the presence of the particular cell or tissue mRNA in a sample using, e.g., in situ hybridization. In other exemplary embodiments, the polynucleotides are used in diagnostics as expressed sequence tags for identifying expressed genes or, as well known in the art and exemplified by Vollrath et al., Science 258:52-59 (1992), as expressed sequence tags for physical mapping of the human genome. The polypeptides according to the invention can be used in a variety of conventional procedures and methods that are currently applied to other proteins. For example, a polypeptide of the invention can be used to generate an antibody that specifically binds the polypeptide. Such antibodies, particularly monoclonal antibodies, are useful for detecting or quantitating the polypeptide in tissue. The polypeptides of the invention can also be used as molecular weight markers, and as a food supplement. Methods are also provided for preventing, treating, or ameliorating a medical condition which comprises the step of administering to a mammalian subject a therapeutically effective amount of a composition comprising a peptide of the present invention and a pharmaceutically acceptable carrier. In particular, the polypeptides and polynucleotides of the invention can be utilized, for example, as potential therapeutics in the treatment of breast, prostate, colon and other cancers. CEA-like polypeptide could also be useful in treating disorders relating to inflammation and autoimmunity. CEA-like polypeptide may also be used as immunosuppressant in organ transplant patients. Also, a soluble CEA-like molecule could serve as a decoy receptor in certain bacterial and viral infections. The methods of the invention also provides methods for the treatment of disorders as recited herein which comprise the administration of a therapeutically effective amount of a composition comprising a polynucleotide or polypeptide of the invention and a pharmaceutically acceptable carrier to a mammalian subject exhibiting symptoms or tendencies related to disorders as recited herein. In addition, the invention encompasses methods for treating diseases or disorders as recited herein comprising the step of administering a composition comprising compounds and other substances that modulate the overall activity of the target gene products and a pharmaceutically acceptable carrier. Compounds and other substances can effect such modulation either on the level of target gene/protein expression or target protein activity. Specifically, methods are provided for preventing, treating or ameliorating a medical condition, including viral diseases, which comprises administering to a mammalian subject, including but not limited to humans, a therapeutically effective amount of a composition comprising a polypeptide of the invention or a therapeutically effective amount of a composition comprising a binding partner of (e.g., antibody specifically reactive for) CEA-like polypeptides of the invention. The mechanics of the particular condition or pathology will dictate whether the polypeptides of the invention or binding partners (or inhibitors) of these would be beneficial to the individual in need of treatment. According to this method, polypeptides of the invention can be administered to produce an in vitro or in vivo inhibition of cellular function. A polypeptide of the invention can be administered in vivo alone or as an adjunct to other therapies. Conversely, protein or other active ingredients of the present invention may be included in formulations of a particular agent to minimize side effects of such an agent. The invention further provides methods for manufacturing medicaments useful in the above-described methods. The present invention further relates to methods for detecting the presence of the polynucleotides or polypeptides of the invention in a sample (e.g., tissue or sample). Such methods can, for example, be utilized as part of prognostic and diagnostic evaluation of disorders as recited herein and for the identification of subjects exhibiting a predisposition to such conditions. The invention provides a method for detecting a polypeptide of the invention in a sample comprising contacting the sample with a compound that binds to and forms a complex with the polypeptide under conditions and for a period sufficient to form the complex and detecting formation of the complex, so that if a complex is formed, the polypeptide is detected. The invention also provides kits comprising polynucleotide probes and/or monoclonal antibodies, and optionally quantitative standards, for carrying out methods of the invention. Furthermore, the invention provides methods for evaluating the efficacy of drugs, and monitoring the progress of patients, involved in clinical trials for the treatment of disorders as recited above. The invention also provides methods for the identification of compounds that modulate (i.e., increase or decrease) the expression or activity of the polynucleotides and/or polypeptides of the invention. Such methods can be utilized, for example, for the identification of compounds that can ameliorate symptoms of disorders as recited herein. Such methods can include, but are not limited to, assays for identifying compounds and other substances that interact with (e.g., bind to) the polypeptides of the invention. The invention provides a method for identifying a compound that binds to the polypeptide of the present invention comprising contacting the compound with the polypeptide under conditions and for a time sufficient to form a polypeptide/compound complex and detecting the complex, so that if the polypeptide/compound complex is detected, a compound that binds to the polypeptide is identified. Also provided is a method for identifying a compound that binds to the polypeptide comprising contacting the compound with the polypeptide in a cell for a time sufficient to form a polypeptide/compound complex wherein the complex drives expression of a reporter gene sequence in the cell and detecting the complex by detecting reporter gene sequence expression so that if the polypeptide/compound complex is detected a compound that binds to the polypeptide is identified.

Method of obtaining peptides with tissue-specific activity and pharmaceutical compositions on their basis

The invention refers to the field of chemistry and concerns the method of obtaining peptides with tissue-specific activity by targeted chemical synthesis. This invention can be employed in medicine to obtain peptide-based pharmaceuticals normalising the functions of various organs and functions. The method of obtaining peptides proposed in this patent claim embraces quantitative amino acid analysis of acetic extracts from tissues, selection on its basis of two amino acids (Glu and Asp) prevailing in the studied tissue, synthesis of the central link from these amino acids and attachment to its N- and C-ends of the amino acids prevailing among the remaining amino acids in the studied tissue. The peptides obtained by the claimed method possess a tissue-specific activity. There is proposed a pharmaceutical composition possessing a tissue-specific activity and containing as its active base one of the peptides obtained by the claimed method or its salts and a pharmaceutically admissible carrier.

1. A method of obtaining a peptide with tissue-specific activity comprising amino acid analysis of an acetic extract from an animal tissue, selection of amino acids prevailing in the studied tissue, synthesis of a central link of the peptide and attachment to its both ends of amino acids prevailing among the remaining amino acids in the studied tissue. 2. The method of claim 1, wherein the amino acid analysis is conducted on an acetic extract from epiphysis tissue. 3. The method of claim 1, wherein the amino acid analysis is conducted on an acetic extract from cerebral cortex tissue. 4. The method of claim 1, wherein the amino acid analysis is conducted on an acetic extract from liver tissue. 5. The method of claim 1, wherein the central link is presented by glutamic (Glu) and aspartic acids (Asp). 6. The method of claim 1, wherein the amino acids are joined to the central link a N- and C-ends. 7. A pharmaceutical composition possessing a tissue-specific activity and containing an active base and a pharmaceutically admissible carrier, wherein the composition contains as its active base an effective quantity of a peptide obtained by the method of claim 1. 8. The composition of claim 7, which contains a salt of the amino group, of carboxyl group, or a salt of an organic or inorganic cation. 9. A pharmaceutical composition possessing a tissue-specific activity and containing an active base and a pharmaceutically admissible carrier, wherein the composition contains as its active base an effective quantity of a peptide obtained by the method of claim 2. 10. A pharmaceutical composition possessing a tissue-specific activity and containing an active base and a pharmaceutically admissible carrier, wherein the composition contains as its active base an effective quantity of a peptide obtained by the method of claim 3. 11. A pharmaceutical composition possessing a tissue-specific activity and containing an active base and a pharmaceutically admissible carrier, wherein the composition contains as its active base an effective quantity of a peptide obtained by the method of claim 4. 12. A pharmaceutical composition possessing a tissue-specific activity and containing an active base and a pharmaceutically admissible carrier, wherein the composition contains as its active base an effective quantity of a peptide obtained by the method of claim 5. 13. A pharmaceutical composition possessing a tissue-specific activity and containing an active base and a pharmaceutically admissible carrier, wherein the composition contains as its active base an effective quantity of a peptide obtained by the method of claim 6. 14. The composition of claim 9, which contains a salt of the amino group, of carboxyl group, or a salt of an organic or inorganic cation. 15. The composition of claim 10, which contains a salt of the amino group, of carboxyl group, or a salt of an organic or inorganic cation. 16. The composition of claim 11, which contains a salt of the amino group, of carboxyl group, or a salt of an organic or inorganic cation. 17. The composition of claim 12, which contains a salt of the amino group, of carboxyl group, or a salt of an organic or inorganic cation. 18. The composition of claim 13, which contains a salt of the amino group, of carboxyl group, or a salt of an organic or inorganic cation.

<SOH> BACKGROUND ON THE INVENTION <EOH>There are known the methods of obtaining complex peptide substances with tissue-specific activity: Thymalin (1), Epithalamin (2), Prostatilen (3), Cortexin (4) and Retinalamin (5). These substances are complexes of low-molecular polypeptides obtained from animal organs and tissues by extraction in 3% acetic add with chlorous zinc and further treatment of the supraprecipitation fluid with acetone (6). The given method of obtaining peptide substances is marked by considerable variability of physical and chemical properties of the exacted peptides and presence of ballast components in them. Limited reserves of required organic raw material and high labour and energy consumption by production are the drawbacks of the said method of obtaining complex peptide substances, which impede their industrial production. There are known the methods of obtaining peptides by classical peptide synthesis in a solution (7) on the basis of active fractions isolated from Thymalin—Glu-Trp (Thymogen) (8), Thymopoietin II—Arg-Lys-Asp-Val-Tyr (Thymopentin) (9), Splenin—Arg-Lys-Glu-Val-Tyr (Splenopentin) (10), Immunoglobulin G—Thr-Lys-Pro-Arg (Taphcin) (11) and other. However, the substances obtained by the given method primarily possess a singly directed spectrum of biological activity (immunoregulatory), instability of the substances in a solution and high doses of application. There are known methods of obtaining peptides of modified structure by classical peptide synthesis in a solution (7): Arg-α-Asp-Lys-Val-Tyr-Arg (Immunophan) (12) structurally distinguished from Thymopentin by the presence of amino acid substitutes with the chain elongated by end Arginine; γ-Glu-Trp (Bestim) (13) structurally distinguished from Thymogen by γ-bond. These substances are also characterised by their narrowly directed immunobiological activity and complicated procedure of chemical synthesis. There is also known a synthetic polymer (Cop 1) inhibiting cellular immune response, which is obtained by chemical synthesis and contains amino acids L-Ala, L-Glu, L-Lys, L-Tyr in the following molar correlation: 6.0:1.9:4.7:1.0 (14). This method of obtaining active substances is not widespread and concerns only the design of polymer Cop 1.

Packaging of a microchip device

A method of packaging a microchip device, an interposer for packaging, and a packaged microchip device. An interposer (7) is placed on microchip devices (1). The interposer (7) comprises an aperture (11) which extends from the interposer surface where external electrical contacts (9) are located to the surface of the microchip devices (1). Electrical contacts (3) on the microchip device surface are accessible through the aperture (11) in order to electrically connect the electrical contacts (3) with the external electrical contacts (9) of the interposer (7). The aperture (11) is divided into at least two openings or aperture regions, separated by a bridge (18). This facilitates the handling of the interposer (7).

1. A method of packaging a microchip device (1), said method comprising: providing a microchip device (1) having a plurality of first electrical contacts (3); providing an interposer (7) with a plurality of second electrical contacts (9) on an outer side of the interposer (7) and with an aperture (11) extending from the outer side through the interposer (7), wherein the aperture (11) is divided into at least two openings (15, 16, 17) by a bridge (18) which connects opposite aperture edges (13); arranging the interposer (7) adjacent to the microchip device (1) so that the first electrical contacts (3) are accessible from the outer side through at least a first of the openings (15, 16, 17); and making electrical connections (5) between respective ones of the first electrical contacts (3) and the second electrical contacts (9). 2. A method according to claim 1, wherein, after making the electrical connections (5), at least the first opening (15) is filled with electrically insulating material by transfer moulding. 3. A method according to claim 1 or 2, wherein at least one of the electrical connections (5) is connected so as to extend from the microchip device (1) through one of the openings (15) and to further extend outside of the opening (15) on the outer side of the interposer (7) and wherein the connection (5) is encapsulated at least outside the opening (15) with insulating material (25) by transfer moulding. 4. A method according to one of claims 1 to 3, wherein the first opening (15) defines a connecting area to be used for locating the electrical connections (5) and wherein the bridge (18) and the nearest one of the electrical connections (5) in the connecting area are located so that their distance is smaller than three times, preferably two times, the average distance between the connections (5) in the connecting area. 5. An interposer (7) for packaging a microchip device (1), comprising: a plurality of electrical contacts (9) on an outer side of the interposer (7), for electrically contacting the packaged microchip device and to be electrically connected with the microchip device (1), an aperture (11) extending from the outer side into the interposer (7), wherein the aperture (11) is divided into at least two openings (15, 16, 17) by a bridge (18) which connects opposite aperture edges (13) and wherein at least a first of the openings extends from the outer side through the interposer (7) in order to allow connection to the microchip device (1) to be made. 6. An interposer according to claim 5, wherein at least the first opening (15, 16) is a frame or window like opening. 7. An interposer according to claim 5 or 6, wherein at least one of the openings which extends from the outer side through the interposer has an open side where the edge defining the opening's circumference is open. 8. A packaged microchip device, comprising: a microchip device (1) having a plurality of first electrical contacts (3); an interposer (7) with a plurality of second electrical contacts (9), with an aperture (11) extending from an outer side through the interposer (7) wherein the aperture (11) is divided into at least two openings (15, 16, 17) by a bridge (18); and electrical conductors (5) which electrically connect the first electrical contacts (3) with corresponding ones of the second electrical contacts (9); wherein the interposer (7) is attached to the microchip device (1); at least one of the conductors (5) extends within the aperture (11); and the aperture (11) and the separate opening (16, 17) are at least partly filled with an electrically insulating material (25) and thereby the at least one conductor (5) is fixed to the interposer (7). 9. A device according to claim 8 with a plurality of packaged microchips wherein the outlines of the microchips (1) each define a package area in which the interposer (7) covers the surface of the corresponding microchip (1), except for the package area regions of the aperture (11); and wherein the bridge (18) extends substantially parallel to the microchip surface from the outside of one of the package areas into this package area. 10. A device according to claim 9, wherein the bridge (18) connects opposite aperture edges located in different package areas.

Cover for golf club protection

A head cover for protection a golf club and a shaft, more particularly to a cover for golf club, of which the golf club can be inserted into or taken out through the opening part of the head cover, then the upper side is open, so that the club can be inserted into or taken out and connecting ribs are formed as one with plastic supporting ribs like umbrella ribs for obtaining the enough space, then after cover sheets are formed inside and outside of the supporting ribs, they are combined with the head cover, which is open upside and then a protection pipe is combined with the connecting ribs.

1. A head cover for protection a golf club wherein its improvement comprising, a golf club which can be inserted into or taken out through the opening part of the head cover. 2. A head cover for protection a golf club as claimed in claim 1 wherein, a few of supporting ribs that are spread upward and the connecting ribs that are extended downward are combined with a protection pipe. 3. A head cover for protection a golf club as claimed in claim 1 and 2 wherein, the cover sheets are formed inside and outside of the supporting ribs. 4. A head cover for protection a golf club as claimed in claim 2 wherein, the above supporting ribs is formed as a barrel. 5. A head cover for protection a golf club as claimed in claim 1 wherein, the elasticity frame, which has an elasticity power, is formed on the boundary of the opening part in the head cover. 6. A head cover for protection a golf club as claimed in claim 1 wherein, after the cover sheets is formed and combined with the protection pipe (3), the ornamental cover (8) is formed. 7. A head cover for protection a golf club as claimed in claim 1 wherein, the sleeve, which has a wing that becomes narrow, is formed on the connecting ribs. 8. A head cover for protection a golf club as claimed in claim 5 wherein, the center opening part of the elasticity frame is open and closed using a string or a patch.

<SOH> BACKGROUND OF THE INVENTION <EOH>1. Field of the Invention The present invention relates to a head cover for protecting a golf club and a shaft, more particularly to a cover for golf club, of which the golf club can be inserted into or taken out through the opening part of the head cover. Usually the golf club is kept in a bag that is divided into two or three equal parts by a partition, so the golf clubs are moving in a disordered way, which causes tangling among the golf clubs and makes inconvenient for inserting into or taking out and causes damage to the shaft and the grip of the golf clubs. Therefore, in order to protect the golf clubs, usually a cover, which is produced with textiles that have a spandex quality, protects by covering the head. However, this kind of cover is inconvenient to use, because it is covered form upside and after covering, it should be pulled out. In addition, when the clubs are not placed vertically in the bottom, they are to be tangled with each other, because the grip's diameter is usually bigger than the shaft's, 2. Discussion of Related Art Thus, the present invention is designed in order to solve the above-mentioned problem, the upper side is open, so that the club can be inserted into or taken out and connecting ribs are formed as one with plastic supporting ribs like umbrella ribs for obtaining the enough space. And after cover sheets are formed inside and outside of the supporting ribs, they are combined with the head cover, which is open upside and then a protecting pipe is combined with the connecting ribs.

<SOH> BRIEF DESCRIPTION OF THE ATTACHED DRAWINGS <EOH>FIG. 1 is a sectional view showing that a protecting pipe is combined with the connecting ribs which are formed supporting ribs; FIG. 2 is a sectional view showing another embodiment of a cover sheet; FIG. 3 is a sectional view showing a state attaching an ornamental cover; FIG. 4 is a state view showing that the cover sheet covers a supporting ribs; FIG. 5 is a perspective view showing that a barrel is used instead supporting ribs; FIG. 6 is a state view showing that the cover sheet covers the barrel; FIG. 7 is a sectional view showing of the embodiment of the present invention used. detailed-description description="Detailed Description" end="lead"?

Multilayered packaging for greasy products

The invention relates to a multilayered packaging for greasy products or part of said packaging, comprising a carrier layer made of a polymer material as main component and at least one layer placed on said carrier layer that does not form the outer side of the packaging, said layer containing a starch derivative as main component. The invention is characterized in the starch derivative is a starch derivative modified with a C2-C6-alkylene oxide. The invention also relates to the use of a C2-C6-alkylene oxide derivatized starch as main component of a layer of a multilayered packaging, which is placed on a carrier layer made of a polymer material in said packaging with the aim of rendering said multilayered packaging grease-tight.

1. Multilayer packaging for greasy products or part of said packaging, comprising a carrier layer made of a polymer material as the main component, and at least one layer applied onto said carrier layer and not forming the outer side of said packaging, said layer containing a starch derivative as the main component, characterized in that said starch derivative is a starch derivative modified with a C2-C6 alkylene oxide 2. Multilayer packaging in accordance with claim 1, characterized in that said starch derivative is a starch derivative modified with a C2-C4 alkylene oxide. 3. Multilayer packaging or part of said packaging in accordance with claim 1, characterized in that said C2-C6 alkylene oxide is propylene oxide. 4. Multilayer packaging or part of said packaging in accordance with any of the aforementioned claims, characterized in that said starch derivative was obtained by modification of corn, wheat, potato, HA pea, or tapioca starch which was optionally partly degraded. 5. Multilayer packaging or part of said packaging in accordance with any of the aforementioned claims, characterized in that said starch derivative has a degree of derivatization of from 0.1 to 1, more preferably of from 0.1 to 0.4. 6. Multilayer packaging or part of said packaging in accordance with any of the aforementioned claims, characterized in that the polymer material of said carrier layer is a polymer occurring in nature, preferably cellulose. 7. Multilayer packaging or part of said packaging in accordance with any of the aforementioned claims, characterized in that said layer containing a starch derivative as the main component contains additional components selected from among pigments, softeners, agents increasing the long-term stability, agents increasing the water stability, and agents affecting the elasticity. 8. Multilayer packaging or part of said packaging, preferably in accordance with claim 4, characterized in that additional components are selected from among glycerol, urea, borax or glyoxal. 9. Use of a starch derivatized with a C2-C6 alkylene oxide as the main component of a layer of a multilayer packaging, said layer being applied onto a carrier layer of said packaging which is made of a polymer material, with the aim of rendering said multilayer packaging greaseproof. 10. Use in accordance with claim 9, characterized in that said C2-C6 alkylene oxide is propylene oxide. 11. Use in accordance with any of claims 9 and 10, characterized in that said starch derivative was obtained by modification of optionally partly degraded corn, wheat, potato, HA pea, or tapioka starch and optionally has a degree of derivatization of from 0.1 to 1, more preferably of from 0.1 to 0.4. 12. Use in accordance with any of claims 9 to 11, characterized in that said layer contains additional components selected from among pigments, softeners, agents increasing the long-term stability, agents increasing the water stability, agents increasing the KIT value, and agents affecting the elasticity, preferably selected from among glycerol, urea, borax, or glyoxal.

<SOH> BACKGROUND OF THE INVENTION <EOH>1. Field of the Invention The invention relates to multilayer greaseproof packaging materials having a carrier layer which is made of paper/cardboard or other suitable substances on a polymer basis. 2. Description of the Related Art It has been known for a long time to provide paper and cardboard containers with coatings having a barrier effect for flavors or humidity/liquids. DE 41 09 983 A1 describes a flexible packaging container comprising a composite made of a paper layer and a thermoplastic layer or film. The thermoplastic layer or film material consists of starch, a synthetic, non-polyolefinic polymer containing hydroxyl groups, e. g. an oxygenated polymer, and softeners of natural origin, e. g. polyalcohols derived from starch. Said material can be melted by supplying heat and therefore, it is extrudable. DE 41 37 802 A1 proposes to laminate a cardboard with a coated paper web to obtain a rottable, liquid-repellent product. The coating of the paper web shall be effected on a starch basis, e. g. on potato starch basis. DE 42 94 110 discloses a coating dispersion which is prepared from copolymers of oxidized starch and styrene, butadiene, acrylic acid or similar polymerizable molecules. Said dispersion reduces the gas and water permeability of cardboard or paper. However, it is often necessary to provide packaging materials which have a high resistance to grease. Thus, food for animals, bread and confectionery, sweets and chocolate require a packaging having a particularly high resistance to grease which is for example indicated by the so-called KIT numbers exhibiting values between 8 and 12. High KIT numbers represent high resistances to grease, values as from 6 already represent a good resistance to grease. Corresponding commercially offered paper/cardboard packaging has usually been subjected to a grease-repellent surface and/or mass treatment. At present, mainly fluoropolymers are used for said mass treatment or surface treatment, up to 5% by weight of coating material being applied onto the material. Grease resistances of >6 to 8 can only be obtained by combining layer and mass treatment, grease resistances with KIT numbers of >12 cannot be guaranteed with the present systems. For example, packing dry food for animals with a low grease content (<10%) requires a mass treatment of the backside, in case of higher grease contents, a barrier is realized by mass treatment in combination with a surface coating. Waste paper, paperboard and cardboard packaging are regularly disposed via the waste paper circuit. Thus, via the pulping process, the halogen polymers used as grease barrier either arrive at the virgin paper product or in the waste water of the process. Starch ethers are known as auxiliary agents and starting materials in the paper industry. The properties used are described in detail in the pertinent literature. They are used in surface coating and coating, respectively, and in pigmented paper coatings. In accordance with the BGVV (Bundesinstitut für gesundheitlichen Verbraucherschutz und Veterinärmedizin), paper, cardboard, and paperboard admitted for food contact may also contain starch ethers (e. g. hydroxyethyl ether and hydroxypropyl ether). Further, starch ethers are used as a component of adhesives because of their good film-forming property and their water bonding capacity. Respective literature is to be found for example in Ullmanns Enzyklopäidie der technischen Chemie; W. Baumann/B. Herberg: Papierchemikalien—Fakten zum Umweltschutz (Springer-Verlag); O. B: Wüirzburg: Modified Starches: Properties and Uses (CRC Press). Further, it is known that starch ether derivatives can be processed to foils/films from an aqueous solution, particularly using casting technology. When preparing the starch ethers in accordance with the so-called Slurry method, the aqueous starch suspension is derivatized under alkaline conditions at temperatures of up to 50° C. The degree of substitution (DS) is substantially around 0.2. The preferred derivatization at the C 2 atom is characteristic for said methods. Another method which is substantially known from scientific examinations (autoclave method) is based on alkaline-activated starch and realizes more homogeneous derivatizations at lower TS (i. e. dry substance) concentrations, the degree of substitution (DS), however, being adjusted similarly. Proceeding in accordance with said strategy is described in DE 42 23 471 A1, the starch ethers obtained according to this document being intended to be used for the preparation of films, particularly for use as overhead, copying, and printing films or for the surface finishing of special papers, and as packaging material. Further, it is indicated in said publication that the ether derivative films mentioned therein can be used in combination with other materials.

<SOH> SUMMARY OF THE INVENTION <EOH>The object of the present invention is to provide packaging materials which are admissible according to food law, and which are greaseproof. Surprisingly, it was found that substrates which themselves do not provide sufficient resistance to grease, such as paper, cardboard, paperboard, or other materials which are made of or contain cellulose, are greaseproof when coated with alkylene-oxide-derivatized starch. Therefore, the present invention provides multilayer packagings for greasy products or parts of said packagings, said packagings comprising a carrier layer made of a polymer material as the main component, and at least one layer applied onto said carrier layer and not forming the outer side of said packaging, said layer applied onto said carrier layer containing an alkyene-oxide-derivatized starch as the main component. The alkylene oxide used for this purpose can suitably be a C 2 -C 6 alkylene oxide. C 2 -C 4 alkylene oxides are preferred. detailed-description description="Detailed Description" end="lead"?

Automotive trim component having an elastomeric skin with a foam core and method for making same

A mold assembly for manufacturing a trim component of an automotive vehicle. The mold assembly comprises a first mold half having a contoured first mold surface and an outer peripheral edge and a second mold half having a contoured second mold surface and an outer peripheral edge. The mold halves pivotal about a hinge between an open position providing access to the respective mold surface and a closed position with the outer peripheral edge of the first mold half aligned with an abutting the outer peripheral edge of the second mold surface. A cutting ridge is formed around the outer peripheral edge of the first mold half for engaging a bulbous projection formed around the outer peripheral edge of the second mold halves in the closed position whereby upon the injection of a urethane material onto the mold halves, the cutting ridge abuts against the projection and perforates any oversprayed material extending outside of the mold surfaces when the mold halves are moved to the closed position.

1. A mold assembly for manufacturing a trim component of an automotive vehicle comprising: a first mold half having a contoured mold surface and an outer peripheral edge; a second mold half having a contoured mold surface and an outer peripheral edge; said mold halves movable between an open position providing access to said respective mold and a closed position with said outer peripheral edge of said first mold half aligned with and abutting said outer peripheral edge of said second mold surface; and a cutting ridge formed around said outer peripheral edge of one of said mold halves for engaging said other of said mold halves in said closed position whereby upon the injection of a urethane material onto at least one of said mold halves said cutting ridge perforates any oversprayed material extending outside of said mold surfaces when said mold halves are moved to said closed position. 2. A mold assembly as set forth in claim 1 further including a projection formed around said outer peripheral edge of said other of said mold halves opposite said cutting ridge for abutting with said cutting ridge with said mold halves in said closed position to thereby cut any oversprayed material therebetween. 3. A mold assembly as set forth in claim 2 further including a mold cavity defined between said mold surfaces of said first and second mold halves in said closed position. 4. A mold assembly as set forth in claim 3 wherein said cutting ridge includes a substantially triangular barb having a sharp leading edge extending upwardly adjacent said outer peripheral edge of said first mold half. 5. A mold assembly as set forth in claim 4 wherein said projection includes a generally bulbous semi-circular configuration extending upwardly adjacent said outer peripheral edge of said second mold half for abutting with said sharp leading edge of said cutting ridge with said mold halves in said closed position. 6. A mold assembly as set forth in claim 5 further including a hinge interconnecting said first and second mold halves for providing pivotal movement of said mold halves between said open and closed positions. 7. A method for manufacturing a trim component for an automotive vehicle with a mold assembly having a first mold half having a first mold surface and a cutting ridge surrounding the first mold surface and a second mold half having a second mold surface and a projection surrounding the second mold surface, wherein the mold halves are moveable relative to each other between an open position having the cutting ridge disengaged from the projection and a closed position having the cutting ridge engage the projection; said method comprising the steps of: applying an in-mold coat to each of the mold halves when the mold halves are in the open position; applying a skin coat to each in-mold coat when the mold halves are in the open position, whereby the skin coat in the first mold half bonds with the respective in-mold coat to form a first structural skin and the skin coat in the second mold half bonds with the respective in-mold coat to form a second structural skin; moving the mold halves to the closed position whereby the cutting ridge abuts the projection to cut any overspray of in-mold coat and skin coat extending outside of the first and second mold surfaces; injecting a foam into the mold halves after applying the skin coats to expand and cure between the first and second structural skins forming the trim component; moving the mold halves to the open position after forming the trim component; and removing the trim component from the mold assembly after moving the mold halves to the open position. 8. A method as set forth in claim 7 further including bonding the first structural skin to the second structural skin by allowing the skins to at least partially harden to a non-liquid state prior to moving the mold halves to the closed position and then abutting the first and second structural skin around the periphery of the first and second mold surfaces. 9. A method as set forth in claim 8 further including venting gas from a mold cavity formed between the mold halves in the closed position as the foam is injected between the first and second structural skins. 10. A method as set forth in claim 9 further including inserting at least one elongated hollow armature between the first and second mold halves in the closed position and injecting the foam into the mold cavity to cure around the armature as part of the trim component. 11. A method as set forth in claim 10 further including injecting the foam through the hollow armature into the mold cavity. 12. A method as set forth in claim 10 further including venting the gas in the mold cavity through the hollow armature during the injecting of the foam between the mold halves. 13. A method as set forth in claim 7 further including injecting the foam into at least one of the mold halves against the in-mold coat and skin coat prior to moving the mold halves to the closed position. 14. A method as set forth in claim 13 further including cutting any overspray of in-mold coat and skin coat and any excess foam outside the periphery of the first and second mold surfaces by abutting the cutting ridge against the projection with the mold halves in the closed position. 15. A method as set forth in claim 7 further including injecting the foam between the first and second structural skins after the mold halves are moved to the closed position and the structural skins are bonded around the periphery of the mold surfaces.

<SOH> BACKGROUND OF THE INVENTION <EOH>1) Field of the Invention The subject invention relates to an automotive trim component, such as a headrest, armrest or the like. Further, the subject invention also relates to a mold assembly and method for manufacturing the aforementioned automotive trim component. 2) Description of the Related Art Automotive trim components, such as headrests and armrests, and their general method of manufacture are known in the art. For example, a conventional method for making headrests includes the following steps. Initially, a plurality of trim cover pieces are manually, either by machine or by hand, sewn together to form a trim cover envelope, also known in the art as an outer shell of the headrest. The outer shell includes sewn seams throughout. Next, the sewn outer shell is positioned into cavity of a mold assembly. Foam is then injected into the sewn outer shell while in the mold assembly to form the headrest. Such conventional methods for making headrests have a number of deficiencies. In particular, upon injecting foam into the sewn outer shell, foam may leak through the sewn seams in the headrest and result in a defective headrest. Further, automotive seating headrests with the sewn seams are prone to tearing, ripping, snagging, and opening during repeated use over the life of an automobile. Illustrative of another example, an alternative method of making a headrest is disclosed in U.S. Pat. No. 5,116,557 wherein the outer shell or layer of the headrest is synthetically made of an elastomer within a mold tool. The mold tool includes a pair of mold halves which form a hermetic seal when closed together. The method as disclosed in the '557 patent requires that the elastomer be in a substantially liquid state during the closing of the mold halves in order to press the elastomer which may have been oversprayed onto the mold halves out from between the mold halves. This method is overly sensitive to the specific curing times of the elastomer and outside factors such as humidity, temperature and the like.