MONOCLONAL ANTIBODIES AND THEIR USE

Abstract: Monoclonal antibodies to the genus Proteus, the labelled antibodies, compositions and kits containing them, and their use in diagnosis of antigen and treatment.

Description: MONOCLONAL ANTIBODIES AND THEIR USE This invention relates to monoclonal antibodies and their use. BACKGROUND OF THE INVENTION Proteus is described in Zinsser Microbiology (17th ed.) 731-2. The organisms of the genus Proteus are found in soil, water, sewage and decaying animal matter, as well as in the human intestinal tract. Brenner's classification scheme recognises two clinically important species in the genus Proteus, Proteus mirabilis and Proteus vulgaris. A third species, Proteus myxofaciens, has been isolated only from gypsy moth larvae. The extreme specificity of antigen-antibody reactions has made it possible to recognise differences between strains of a bacterial species, such as the above-mentioned species, that are indistinguishable on the basis of other phenotypic criteria. . mirabilis accounts for the majority of Proteae infections in man, causing community-acquired, as well as hospital-acquired urinary tract infections.- Wound infections t pneumonia and septicemia can also occur. In addition to urinary tract infections, Proteus is known to cause gram-negative sepsis which is a bloodstream infection. It is one of the major infectious disease problems encountered in modern medical centres. While it can be transient and self-limited, severe gram-negative sepsis constitutes a medical emergency. At the present time, the test for gram-negative sepsis involves processing blood and urine cultures and other procedures on occasion. In addition to being expensive, blood culture tests are cumbersome. They require a day, and often several days, to return results. They require expert laboratory skills because of the complex nature of human blood which tends to interact non-specifically with many of the test reagents. Presently, in urinary tract infections, a microscopic examination is made, to determine the presence of micro-organisms as a preliminary screening. The microscopic examination cannot distinguish among the gram-negative bacteria. Accordingly, a second step is a urine culture to identify the organism isolated in the urine sample. A delay in diagnosis and initiation of treatment can result in serious complications. Thus, existing methods of detection of Proteus with high accuracy in urinary tract infections or gram-negative sepsis are less than satisfactory in that they consume large amounts of expensive skilled labour and laboratory time, generally taking one and often several days before returning results. The production of monoclonal antibodies is now a well-known procedure first described by Kohler and Milstein, Eur. J. Immunol.. (1975) 292. While the general technique of preparing hybridomas and the resultant monoclonal antibodies is understood, it has been found that preparing a specific monoclonal antibody to a specific antigen is difficult, mainly due to the degree of specificity and variations required in producing a particular hybridoma. SUMMARY OF THE INVENTION The present invention provides novel monoclonal antibodies for use in accurately and rapidly diagnosing samples for the presence of Proteus antigens and/or organisms. Briefly stated, the present invention comprises monoclonal antibodies specific for an aritigen of Proteus? in particular, the antigens of Proteus mirabilis and the antigens of Proteus vulgaris, as well as a monoclonal antibody broadly cross-reactive with an antigen for each species of the genus Proteus. The invention also comprises labelled monoclonal antibodies for use in diagnosing the presence of the Proteus antigens, each comprising a monoclonal antibody -3- against one of the above-mentioned antigens to Proteus or to a particular species thereof and having linked thereto an appropriate label. The label can be, for example, a radioactive isotope, enzyme, fluorescent compound, chemiluminescent compound, bioluminescent compound, ferromagnetic atom or particle. The invention further comprises the process for diagnosing the presence of Proteus antigens or organisms in a specimen, comprising contacting said specimen with the labelled monoclonal antibody in an appropriate immunoassay procedure. Additionally, the invention is also directed to a therapeutic composition comprising a monoclonal antibody for an antigen of Proteus and a carrier or diluent, as well as kits containing at least one labelled monoclonal antibody to an antigen of a Proteus. DETAILED DESCRIPTION The monoclonal antibodies of the present invention are prepared by fusing spleen cells from a mammal which has been immunised against the particular Proteus antigen, with an appropriate myeloma cell line, preferably NSO (uncloned) , P3NS1-Ag4/1, or Sp2/0 Agl4. The resultant product is then cultured in a standard HAT (hypoxanthine, a inopterin and thymidine) medium. Screening tests for the specific monoclonal antibodies are employed utilising immunoassay techniques which will be described below. The immunised spleen cells may be derived from any mammal, such as primates, humans, rodents (i.e. mice, rats and rabbits) , bovines, ovines and canines, but the * present invention will be described in connection with mice. The mouse is first immunised by injection of the particular Proteus antigen chosen, generally for a period of approximately eleven weeks. When the mouse shows sufficient antibody production against the antigen, as -4- determined by conventional assay, it is given a booster injection of the appropriate Proteus antigen, and then killed so that the immunised spleen may be removed. The fusion can then be carried out utilising immunised spleen cells and an appropriate myeloma cell line. The fused cells yielding an antibody which gives a positive response to the presence of the particular Proteus antigen are removed and cloned utilising any of the standard methods. The monoclonal antibodies from the clones are then tested against standard antigens to determine their specificity for the particular Proteus antigen ^ The monoclonal antibody selected, which is specific for the particular Proteus antigen or species, is then bound to an appropriate label. Amounts of antibody sufficient for labelling and subsequent commercial production are produced by the known techniques, such as by batch or continuous tissue culture or culture in vivo in mammals such as mice. The monoclonal antibodies may be labelled with various labels, as exemplified above. The present invention will be described with reference to the use of an enzyme-labelled monoclonal antibody. Examples of enzymes utilised as labels are alkaline phosphatase, glucose oxidase, galactosidase, peroxidase and urease. Such linkage with enzymes can be accomplished by any known method, such as the Staphylococcal Protein A method, the glutar ldehyde method, the benzoquinone method, or the periodate method. Once the labelled monoclonal antibody is formed, testing is carried out employing one of a wide variety of conventional immunoassay methods. The particular method chosen will vary according to the monoclonal antibody and the label chosen. At the present time, enzyme immunoassays are preferred owing to their low cost, reagent stability, safety, sensitivity and ease of -5- procedure. One example is the enzyme-linked im unosorbent assay (EIA) . EIA is a solid-phase assay system which is similar in design to the radiometric assay, but which utilises an enzyme in place of a radioactive isotope as the immunoglobin marker. Fluorescent-immunoassay is based on the labelling of antigen or antibody with fluorescent probes. A non-labelled antigen and a specific antibody are combined with identical fluorescently-labelled antigen. Both labelled and unlabelled antigen compete for antibody binding sites. The amount of labelled antigen bound to the antibody is dependent upon, and therefore a measurement of, the concentration of non-labelled antigen. Examples of this particular type of fluorescent-immunoassay include heterogeneous systems such as Enzyme-Linked Fluorescent Immunoassay, or homogeneous systems such as the Substrate-Labelled Fluorescent Immunoassay. The most suitable fluorescent probe r and the one most widely used, is fluorescein. While fluorescein can be subject to considerable interference from scattering, sensitivity can be increased by the use of a fluorometer optimised for the probe utilised in the particular assay, and in which the effect of scattering can be minimised. in fluorescence polarisation, a labelled sample is excited with polarised light and the degree of polarisation of the emitted light is measured. As the antigen binds to the antibody, its rotation slows down and the degree of polarisation increases. Fluorescence polarisation is simple, quick and precise. However, at the present time, its sensitivity is limited to the micromole per litre range and upper nanomole per litre range with respect to antigens in biological samples. Luminescence is the emission of light by an atom or molecule as an electron is transferred to the ground state from a higher energy state. In both chemiluminescent and biolu inescent reactions, the free energy of a chemical reaction provides the energy required to produce an intermediate reaction or product in an electronically-excited state. Subsequent decay back to the ground.state is accompanied by emission of light. Bioluminescence is the name given to a special form of chemiluminescence found in biological systems, in which a catalytic protein or enzyme, such as luciferase, increases the efficiency of the luminescent reaction. The best known chemiluminescent substance is luminol. A further aspect of the present invention is a therapeutic composition comprising one or more of the monoclonal antibodies to the particular Proteus antigen or species, as well as a pharmacologically- acceptable carrier or diluent. Such compositions can be used to treat humans and/or animals afflicted with some form of Proteus infection and they are used in amounts effective to cure; the amount may vary widely, depending upon the individual being treated and the severity of the infection. One or more of the monoclonal antibodies can be assembled into a diagnostic kit for use in diagnosing for the presence of an antigen, antigens or species of Proteus in various specimens. It is also possible to use the broadly cross-reactive monoclonal antibody which can identify the genus Proteus alone or as part of a kit containing antibodies that can identify other bacterial genera or species of Proteus and/or other bacteria. In the past, there have been difficulties in developing rapid kits because of undesirable cross-reactions of specimens; e.g. urine with antiserum. The use of monoclonal antibodies can eliminate these problems and provide highly specific and rapid tests for diagnosis. For example, the incidence of significant diarrhea and diarrheal illness is so high that estimates of market size for such a kit are difficult to make, but a "same day" test could be expected to be used at least as often as stool cultures. Large use of such tests in developing countries might be anticipated because of more frequent and severe diarrhea, and other related illnesses. Additionally, a kit could be used in pathology laboratories for the rapid detection of gram-negative bacteria in urine, or on an out-patient basis. Further, conjugated or labelled monoclonal antibodies for antigens and/or species of Proteus and other gram-negative bacteria can be utilised in a kit to identify such antigens and organisms in blood samples taken from patients for the diagnosis of possible Proteus or other gram-negative sepsis. The monoclonal test is an advance over existing procedures in that it is more accurate than existing tests; it gives "same day" results, provides convenience to the patient and improves therapy as a result of early, accurate diagnosis; and it reduces labour costs and laboratory time required for administration of the tests. The kit may be sold individually or included as a component in a comprehensive line of compatible immunoassay reagents sold to reference laboratories to detect the species and serotypes of Proteus. One preferred embodiment of the present invention is a diagnostic kit comprising at least one labelled monoclonal antibody against a particular Proteus antigen or species, as well as any appropriate stains, counterstains or reagents. Further embodiments include kits containing at least one control sample of a Proteus antigen and/or a cross-reactive labelled monoclonal antibody which would detect the presence of any of the given particular Proteus organisms in a particular sample. Monoclonal diagnostics which detect the presence of Proteus antigens can also be used in periodic testing of water sources, food supplies and food processing operations. Thus, while the present invention describes the use of the labelled monoclonal antibodies to determine the presence of a standard antigen, the invention can have many applications in diagnosing the presence of antigens by determining whether specimens, such as urine, blood, stool, water and milk, contain the particular Proteus antigen. More particularly, the invention could be utilised as a public health and safety diagnostic aid, whereby specimens such as water or food could be tested for possible contamination. The invention will be further illustrated in connection with the following Examples which are set forth for purposes of illustration only and not by way of limitation. The monoclonal * antibodies of the present invention were prepared generally according to the method of Kohler and Milstein, supra. In the Examples: API = Analytical Profile Index (ref. Ayerst Laboratories) DMEM = Dulbeccos Modified Eagles Medium FCS = Foetal Calf Serum % T refers to vaccine concentrations measured in a 1 cm light path PBS = Phosphate Buffered Saline Example 1 A. Antigen Preparation Proteus mirabilis was obtained from the National Collection of Type Cultures (NCTC accession No. 60) and tested against standard reference typing sera to confirm -9- its typing. More specifically, the antigen was removed from the lyophile, grown on blood agar, and tested by conventional biochemical (API) and agglutination tests with appropriate antisera to confirm it identity and purity. The cells were then transferred to DMEM, grown, and harvested for use as a source of antigen. The organisms were washed in formol saline by repeated centrifugation and were resuspended in formol saline. B. Animal Immunisation Six Balb/c mice were injected with the prepared antigen. They were given 3 weekly intraperitoneal injections (0.05 ml of 80% T vaccine) of boiled killed Proteus mirabilis prepared as above, followed by 3 weekly intravenous injections and a further intravenous injection after a month. The mice were bled approximately six days after the last injection .and the serum tested for antibodies by assay. The conventional assay used for this serum titer testing was the enzyme-linked im unosorbent assay system. When the mice showed antibody production after this regimen, generally a positive titer of at least 10,000, a mouse was selected as a fusion donor and given a booster injection (0.02 ml of 80% T vaccine) intravenously, three days prior to splenectomy. c. Cell Fusion The selected donor mouse was killed and surface sterilised by immersion in 70% ethyl alcohol. The spleen was then removed and immersed in approximately 2.5 ml of DMEM to which had been added 3% FCS. The spleen was then gently homogenised in a LUX homogenising tube until all cells had been released from the membrane and the cells were washed in 5 ml 3% FCS DMEM. The cellular debris was then allowed to settle and the spleen cell suspension placed in a 10 ml centrifuge tube. The debris was then rewashed in 5 ml 3% FCS DMEM. 50 ml of suspension were then made in 3% FCS DMEM. The myeloma cell line used was NSO (uncloned) , obtained from the MRC Laboratory of Molecular Biology in Cambridge, England. The myeloma cells were in the log growth phase, and rapidly dividing. Each cell line was washed using a tissue culture medium DMEM containing 3% FCS. The spleen cells were then spun down at the same time that a relevant volume of myeloma cells were spun down (room temperature for 7 minutes at 600 g) , and each resultant pellet was then separately resuspended in 10 ml 3% FCS DMEM. In order to count the myeloma cells, 0.1 ml of the suspension was diluted to 1 ml and a haemacytometer with phase microscope was used. In order to count the spleen cells, 0.1 ml of the suspension was diluted to 1 ml with Methyl Violet-citric acid solution r and a haemacytometer and light microscope were used to count the stained nuclei of the cells. 8 7 1.3 x 10 spleen cells were then mixed with 8 x 10 myeloma cells, the mixture washed in serum-free DMEM high in glucose and centrifuged, and all the liquid removed. The resultant cell pellet was placed in a 37°C water bath. Over the period of one minute, 1 ml of a 50% w/v solution of polyethylene glycol 1500 (PEG) in saline Hepes, pH of approximately 7.5, is added, and the mixture gently stirred for approximately 1.5 minutes. There were then slowly added 10 ml of serum-free tissue culture medium DMEM, followed by the addition of up to 50 ml of such culture medium, centrifugation and removal of all the supernatant, and resuspension of the cell pellet in 10 ml of DMEM containing 18% by weight FCS. 10 μl of the mixture were placed in each of 480 wells of standard multiwell tissue culture plates. Each well contains 1.0 ml of the standard HAT medium (hypoxanthine, aminopterin, and thymidine) and a feeder layer of Balb/c 4 macrophages at a concentration of 5 x 10 macrophages/well. The wells were kept undisturbed and cultured at 37°C in 9% C0 2 -air at approximately 100% humidity. The wells were analysed for growth utilising the conventional inverted microscope procedure, after about 5 to 10 days. In those wells in which growth was present in the inhibiting HAT medium, screening tests for the specific monoclonal antibody were made utilising the conventional enzyme immunoassay screening method described below. Somewhere around 10 days to 14 days after fusion, sufficient antibody against the antigen was developed in at least one well. D. Cloning From those wells which yielded antibody against the antigen, cells were removed and cloned using the standard agar method (twice) and by limiting dilution. In the agar method, a freshly-prepared stock solution of sterile 1.2% agar in double-distilled water with an equal volume of double-strength DMEM and additives was maintained at 45°C. This solution (10 ml) was then aliquoted into 10 cm Petri dishes, to form a base layer. An overlay of equal volumes of agar and cells in 18% FCS-DMEM was spread evenly over the base. The cells were allowed to multiply for approximately 10 days at 37°C, 7-9% CO ™ , 9.5% RH. Viable separate colonies were picked off the agar surface and placed into 60 wells of a 96-well icrotitre tray in 18% FCS-DMEM. After a further period of growth, the supernatants were assayed for specific antibody by the standard enzyme immunosorbent assay. In limiting dilution, dilutions of cell suspensions in 18% FC-DMEM + Balb/c mouse macrophages were made to achieve 1 cell/well and half cell/well in a 96-well microtitre plate. The plates were incubated for 7-14 days at 37°C, 95% RH, 7-9% C0 2 , until semi-confluent. The supernatants were then assayed for specific antibody by the standard enzyme im unosorbent assay. E. Monoclonal Selection The monoclonal antibodies from the clones were screened by the standard techniques for binding to the antigen prepared as in the immunisation, and for specificity in a test battery of Proteus species and related genera bearing different antigens. Specifically, a grid of microtiter plates containing a representative selection of O-serotype organisms, i.e. Providencia, Shigella, Serratia and Salmonella, was prepared, boiled, and utilised as a template to define the specificity of the parent O-specific group. The EIA immunoassay noted above was used. F. Antibody Production Cells of the monoclonal antibody-producing cell line were grown in batch tissue culture. 10% FCS-DMEM was used to support growth in mid-log phase, to a 1 litre volume, then the culture was allowed to overgrow to allow maximum anitbody production. The culture was then centrifuged at 1200 g for approximately 10 min, the cells discarded and the antibody-rich supernatant collected. The fluid was titrated, as noted above, to establish presence and level of antibody, and purified. Purification is accomplished using the supernatant on Protein A-Sepharose method, in which, to 1 litre of culture supernatant were added 100 ml of 1.0M Tris buffer, pH 8.2. ' The tris buffered supernatant was applied at a flow rate of 1 ml/min to a 1 ml column of Protein A-Sepharose, previously equilibrated with 0.1M Tris buffer, pH 8.2. The column was then washed with 40 ml of 0.1M Tris buffer. The monoclonal antibody was eluted with citrate buffer (0.1M sodium citrate, pH 3.5) into sufficient 1M Tris buffer, pH 9.0, to raise the pH immediately to about 7.5. The eluate was dialysed in PBS, pH 7.4, at 4°C, and stored at -20°C. G. Enzyme-Monoclonal Linkage The monoclonal antibody specific against Proteus mirabilis antigen, prepared and screened as described above, is then bound to an appropriate enzyme; in this case, a highly purified alkaline phosphatase. This is accomplished by the benzoquinone conjugation method. 24 mg alkaline phosphatase (Sigma Type VII-T) were dialysed against 2 x 500 ml of 0.25M sodium phosphate buffer, pH 6.0, at +4°C. Para-benzoquinone, 18 mg, was dissolved in warm AR ethanol, 0.6 ml, and added to the dialysed alkaline phosphatase. The benzoquinone/alkaline phosphatase mixture was left in the dark at room temperature for 1 hour. After this time, unreacted benzoquinone and reaction by-products were removed and the buffer exchanged, by gel filtration on a Pharmacia ' PD-10 (Sephadex G-25M) column, previously equilibrated in 0.15M sodium chloride. The benzoquinone-activated alkaline phosphatase thus produced was sufficient for six 1.5 mg antibody conjugations. Monoclonal antibody was dialysed against 2 x 500 ml of 0.15M sodium chloride at +4"C. Dialysed antibody was added to 8 mg of benzoquinone-activated alkaline phosphatase, immediately followed by sufficient 1M sodium bicarbonate to give a final concentration of 0.1M. The conjugation mixture was left in the dark at +4°C for 48 hours. After this time, sufficient 1M lysine was added to give a final concentration of 0.1M. After 2 hours in the dark at room temperature, the conjugate was dialysed against 2 x 1000 ml of PBS + 0.02% sodium azide at +4°C. An equal volume of glycerol was added. The conjugate was sterile- filtered through a 0.22 μm membrane filter into a sterile amber vial and stored at +4°C. H. Monoclonal Antibody Conjugate Testing The enzyme immunoassay method was used for testing. This assay method comprises coating the wells of a standard polyvinyl chloride miσrotiter tray with the antigen, followed by addition of monoclonal antibody enzyme conjugate, and finally addition of the enzyme substrate, para-nitrophenol phosphate. In this case, the monoclonal antibody was found to be specific for the antigen Proteus mirabilis. The monoclonal antibody was also tested and shown to be of the Class IgG3. If deemed necessary, the particular epitopic site to which the antibody attaches to the antigen can also be determined. The same enzyme immunoassay method can also be used to determine whether diagnostic specimens such as urine, blood, stool, water or milk contain the antigen. In such cases, the antibody can first be bound to. the plate. Example 2 The same procedure as in Example 1 is utilized, in preparing the monoclonal antibody conjugate specific for the antigen of Proteus vulgaris, except for the following differences. Proteus vulgaris was obtained from the National Collection of Type Cultures (NCTC accession No. 4175) . In the animal immunisation step, the intraperitoneal injections were given weekly for 3 weeks, followed by intravenous injections for a further 3 weeks. In the g cell fusion step, 1 x 10 spleen cells were mixed with 6 10 myeloma cells. One agar method step was used in cloning. In this case, a monoclonal antibody was obtained and found to be specific for the antigen of Proteus vulgaris. The sub-class was IgG2a. Example 3 The same procedure as in Example 1 may be utilized in preparing a monoclonal antibody broadly cross-reactive with an antigen of many or all species of the genus Proteus, but using another Proteus obtained from the National Collection of Type Cultures. Tests using the present invention are superior to the existing tests based on the following advantages: (i) greater accuracy; (ii) same day results, within an hour or two; (iii) reduction in amount of skilled labor required to administer laboratory procedures, resulting in reduced labor costs; (iv) reduction in laboratory time and space used in connection with tests, resulting in reduced overhead expense; and (v) improved therapy based upon early, precise diagnosis. While the invention has been described in connection with certain preferred embodiments, it is not intended to limit the scope of the invention to the particular form set forth, but, on the contrary, it is intended to cover such alternatives, modifications, and equivalents as may be included within the spirit and scope of the invention as defined by the appended claims.

Claims: -16- HAT IS CLAIMED IS: 1. A monoclonal antibody specific for an antigen or species of Proteus. 2. The antibody of Claim 1 specific to the antigen or antigens of Proteus mirabilis. 3. The antibody of Claim 1 specific to the antigen or antigens of Proteus vulgaris. 4. A monoclonal antibody broadly cross- reactive with an antigen of all species of the genus Proteus. 5. A labeled monoclonal antibody consisting essentially of a monoclonal antibody of Claims 1-4 and an appropriate label. 6. The labeled, monoclonal antibody of Claim 5, wherein said label is a member of the group selected from a radioactive isotope, enzyme, fluorescent compound, bioluminescent compound, chemiluminescent compound, or ferromagnetic atom, or particle. 7. The labeled monoclonal antibody of Clai 6, wherein said label is an enzyme capable of conjugating with a monoclonal antibody and of being used in an enzyme-linked immunoassay procedure. 8. The labeled monoclonal antibody of Claim 7, wherein said enzyme is alkaline phos¬ phatase, glucose oxidase, galactosidase, or peroxidase. 9. The labeled monoclonal antibody of Claim 6, wherein said label is a fluorescent compound or probe capable of being used in an immuno-fluorescent or fluorescent immunoassay procedure, enzyme fluorescent immunoassay, or fluorescence polarization immunoassay, photon counting immunoassay, or the like procedure. 10. The labeled monoclonal antibody of Claim 9, wherein said fluorescent compound or probe is fluorescein. 11. The labeled monoclonal antibody of Claim 6, wherein said label is a chemiluminescent compound capable of being used in a luminescent or enzyme-linked luminescent immunoassay. 12. The labeled monoclonal antibody of Claim 11, wherein such chemiluminescent compound is luminol or a luminol derivative. 13. The labeled monoclonal antibody of Claim 6, wherein said label is a bioluminescent compound capable of being used in an appropriate bioluminescent immunoassay. 14. The labeled monoclonal antibody of Claim 13, wherein such bioluminescent compound is luciferase or a luciferase derivative. 15. A process for diagnosing for the pre¬ sence of an antigen of Proteus in a specimen comprising contacting at least a portion of said specimen with a labeled monoclonal antibody of Claim 5 in an immunoassay procedure appropri¬ ate for said label. 16. The process of Claim 15, wherein the appropriately labeled immunoassay procedure is selected from immuno-fluorescent or fluorescent immunoassay, i muno-electron microscopy, radio- metric assay systems, enzyme-linked immunoassays. fluorescence polarization, photon-counting bio¬ luminescent, or chemiluminescent immunoassay. 17. The process of Claim 16, wherein said label is an enzyme capable of being used in an enzyme-linked immunoassay procedure. 18. The process of Claim 17, wherein said enzyme is selected from alkaline phosphatase, glucose oxidase, galactosidase, or peroxidase. 19. The process of Claim 16, wherein said label is a fluorescent compound or probe capable of being used in an immuno-fluorescent or fluores¬ cent immunoassay procedure, enzyme fluorescent immunoassay, or fluorescence polarization immuno¬ assay, or photon-counting immunoassay, or the like procedure. 20. The process of Claim 19, wherein said fluorescent compound or probe is fluorescein. 21. The process of Claim 16, wherein said label is a chemiluminescent compound capable of being used in a luminescent or enzyme-linked luminescent immunoassay. -20- 22. The process of Claim 21, wherein said chemiluminescent compound is luminol or a luminol derivative. 23. The process of Claim 16, wherein said label is a bioluminescent compound capable of being used in a bioluminescent or enzyme-linked bioluminescent immunoassay. 24. The process of Claim 23, wherein said bioluminescent compound is luciferase or a lucif- erase derivative. 25. A therapeutic composition comprising one or more of the labeled monoclonal antibodies in Claims 1-4 and a pharmaceutically acceptable carrier or diluent. 26. A therapeutic composition comprising one or more of the labeled monoclonal antibodies in Claim 5 and a pharmaceutically acceptable carrier or diluent. 27. A method of treating Proteus infec¬ tions comprising administering an effective amount of a monoclonal antibody of Claims 1-4. 28. A kit for diagnosing for the presence of an antigen or species of Proteus in a diagnos¬ tic specimen comprising at least one monoclonal antibody of Claims 1-4. 29. The kit of Claim 28, wherein said at least one antibody is labeled. 30. The kit of Claim 29, wherein said at least one monoclonal antibody is labeled with a fluorescent compound. 31. The kit as in Claim 29, wherein said at least one monoclonal antibody is labeled with an enzyme. 32. The kit as in Claim 29, wherein said at least one monoclonal antibody is labeled with a member of the group consisting of a radio¬ active isotope, chemiluminescent compound, bio¬ luminescent compound, ferromagnetic atom, or particle. 33. The kit of Claims 29, 30, 31, and 32 additionally containing at least one known Proteus antigen as a control. 34. The kit of Claims 29, 30, 31, 32, and 33 containing each known antigen of Proteus mirabilis, and Proteus vulgaris. 35. The kit of Claims 29, 30, 31, 32, and 33 containing the antigens of Proteus mirabilis. 36. The kit of Claims 29, 30, 31, 32, and 33 containing the antigens of Proteus vul¬ garis. 37. A kit for diagnosing for the presence of an antigen or species of Proteus in a diagnos¬ tic specimen comprising at least one monoclonal antibody of Claims 1-4 and a control. 38. The kit of Claim 37, wherein said at least one antigen is labeled and said control is at least one known antigen of Proteus. 39. A kit for diagnosing for the presence of a gram-negative bacterial infection comprising at least one πionoclonal antibody of Claims 1-4. -23- 40. The kit of Claim 39, wherein said at least one monoclonal antibody is labeled.

Inventors: COX PETER JOHN, NOYES ALICE MARGARET, WIDDOWS DANNY, WRIGHT BRUCE WILLIAM, COX, PETER, JOHN, NOYES, ALICE, MARGARET, WIDDOWS, DANNY, WRIGHT, BRUCE, WILLIAM
Applicants: TECHNOLOGY LICENCE CO LTD
Application year: 1985
Main classification: C12P21
IPC codes: A61K38, A61K39, C07K1, C07K14, C07K16, C07K19, C12N5, C12N15, C12P21, C12Q1, C12R1, G01N33