bluuebunny/crossref_metadata_2025_split huggingface dataset:
Novel poly‐substituted aryl acridinium esters and their use in immunoassay
Authors: Law Say‐Jong , Miller Thomas , Piran Uri , Klukas Carol , Chang Steve , Unger John 
Published: May 2005

AbstractA new series of stable acridinium ester conjugates have been developed for use as non‐isotopic labels in immunoassay. They have proved to be a flexible alternative to radioimmunoassay. We present data showing the successful development of immunoassays in sandwich, competitive and receptor formats. In addition, hydrophilic acridinium ester analogues have been synthesized, encapsulated in liposomes, and utilized as labels in immunoassay. The potential of this technology is discussed.
Luciferin derivatives in bioluminescence‐enhanced enzyme immunoassays
Authors: Miska Werner , Geiger Reinhard 
Published: May 2005

AbstractUltrasensitive bioluminescence immunoassays for the determination of peptides and proteins (illustrated with human urinary kallikrein, bradykinin and the determination of human urinary kallikrein antibody titres) have been developed. The usable ranges of the standard curves are from 5 pg to 5000 pg per litre. The relative intra‐assay coefficients of variation of the tests were between 2% and 6%, and the inter‐assay coefficients of variation between 4% and 12%.
T4 and ultrasensitive TSH immunoassays using luminescent enhanced xanthine oxidase assay
Authors: Baret A. , Fert V. 
Published: May 2005

AbstractThe use of xanthine oxidase in immunoanalysis has never been reported. We describe here a procedure in which the xanthine oxidase dependent luminescence of luminol is enhanced in the presence of Fe–EDTA complex, providing an highly sensitive assay (3 amol of enzyme) and a long‐term signal.This specific amplification has been applied to T4 and ultrasensitive TSH solid phase immunoassays, with T4–XO and anti‐TSH monoclonal antibody‐XO conjugates as tracers. The performances of these assays are at least equivalent to those obtained with iodinated tracers, using the same solid phases and the same calibrators. The major advantages of these immunoassays are: (1) the long‐term signal which can be repeatedly recorded over several days, (2) the high detection sensitivity, (3) the long‐term stability of the luminescence reagent and (4) the stability of the conjugates.
Luminescent immunoassay using isoluminol derivatives
Authors: Messeri Gianni , Orlandini Alessandro , Pazzagli Mario 
Published: May 2005

AbstractIsoluminol derivatives (ID) were early employed in the preparation of tracers for immunoassay. Their wide use was mainly due to their high quantum efficiency, low molecular weight, well‐known chemical structure, low cost and high stability. Moreover the light efficiency of some ID may be modified by specific binding to the antibody, thus allowing the development of homogenous immunoassays requiring no bound/free separation step. Some ID are commercially available under both the amino terminal and carboxyl terminal form for conjugation to respectively carboxy derivative of steroid and amino residues of protein. The linking reaction can be commonly carried on via active esters chemistry and can be easily accomplished within one day. Steroid conjugates can then be rapidly purified by silica gel thin layer chromatography, and protein conjugates by gel filtration on a short disposable column. In the field of steroid studies, isoluminol derivative conjugates were prepared for the immunoassay of almost all compounds of clinical interest. When dealing with protein, both antigens and antibody were labelled in this way, resulting in highly specific activity tracers for competitive and non‐competitive immunoassays. Recently the labelling of streptavidin with amino‐buty‐ethyl‐isoluminol allowed the development of very sensitive immunoassay methods which take advantage of the biotin‐avidin system.
An investigation on the catalytic mechanism of enhanced chemiluminescence: Immunochemical applications of this reaction
Authors: Vlasenko S. B. , Arefyev A. A. , Klimov A. D. , Kim B. B. , Gorovits E. L. , Osipov A. P. , Gavrilova E. M. , Yegorov A. M. 
Published: May 2005

AbstractThe mechanism of peroxidase‐catalysed oxidation of luminol by H2O2 was studied. The stopped‐flow technique was used to measure the rate constants for the reactions between the oxidized forms of peroxidase with luminol and the following substrates: p‐iodophenol, p‐bromophenol, p‐clorophenol, o‐iodophenol, m‐iodophenol, luciferin, and 2‐iodo‐6‐hydroxybenzothiazole. The correlation between kinetic parameters and the degree of enhancement was established. The effect of charged synthetic polymers and specific antibodies on the peroxidase activity in the enhanced chemiluminescent reaction. Novel homogenous methods of luminescent immunoassay (LIA) for (1) antibodies to insulin, (2) insulin and (3) antibodies to trinitrophenyl group are proposed on the basis of regulatory facilities of the enhanced chemiluminescent reaction. Based on the enhanced chemiluminescent reaction a peroxidase flow‐injection assay was developed and successfuly tested in the flow‐injection enzyme immunoassays for human IgG and for thyroxin (T4). The immunoassay proposed has a detection limit of 10−9M for IgG and 10−11M for T4, the overall time of the assay being 5–15 min.
Near infrared emission of singlet oxygen generated in the dark
Authors: Khan Ahsan U. 
Published: May 2005

AbstractSinglet oxygen generation is reported from (1) enzymatic reaction and (2) electron transfer reactions of the superoxide anion measured directly with an ultrasensitive near‐IR emission spectrophotometer by monitoring the O2(1Δg) → O2(3Σg−) transition at 1268 nm. Near‐IR emission spectra from the myeloperoxidase and lactoperoxidase enzymatic systems show only emission of singlet oxygen at 1268nm. The lipoxygenase/Na–linoleate enzymatic reaction exhibits two emissions, 1268 nm and 1288 nm. The latter emission is identified as originating from a peroxy radical. Spectral and kinetic data giving evidence of singlet oxygen generation is obtained from the reaction of potassium superoxide solubilized by 18‐crown‐6‐ether in acetonitrile with a series of organometallic coordination compounds.
Hydroxycoumarins as sensitizers and reactants of chemiluminescent oxidative reactions
Authors: Slawinska Danuta , Slawinski Janusz 
Published: May 2005

AbstractChemiluminescence (CL) of 7‐hydroxycoumarin, umbelliferone, a strongly fluorescing compound, the precursor of photoactive furocoumarins widely spread in plant kingdom and used in dye lasers has been examined in four oxidative systems producing active oxygen species: A–horseradish peroxidase + H2O2 + buffers pH = 3–10; BK3Fe (CN)6 + H2O2 + buffers pH = 4–12; CNaOCI + buffer pH = 3–10.8; and DHCHO + H2O2 + K2CO3 (Na2CO3), the so‐called Trautz–Schorigin reaction. In all these systems a 10–1000‐fold increase of CL intensity and quantum yield and an appearance of the blue emission band (λmax = 460–480 nm), typical of the fluorescence of hydroxycoumarin used have been observed. Spectrophotometric and fluorimetric measurements of the systems A–D indicate that the tested hydroxycoumarins ‐ coumarin, umbelliferone and 6‐glucoside umbelliferone or esculin – undergo slow (systems A and B) and fast (systems C and D) consumption, leading to the degradation of coumarins. These findings suggest that efficiently fluorescing hydroxy‐coumarins may act both as sensitizers – secondary emitters of CL and/or reactants undergoing oxidative degradation.
Oxidative stress in the rat heart, studies on low‐level chemiluminescence
Authors: Ursini Fulvio , Barsacchi Renata , Pelosi Gualtiero , Benassi Antonio 
Published: May 2005

AbstractDetection of ultraweak chemiluminescence (CL) emission from the surface of the organ is a sensitive and non‐disruptive tool to evaluate the oxidative stress in rat heart. Indeed, an increased photon emission rate can be observed when cellular antioxidants such as glutathione or vitamin E are depleted, or when organic hydroperoxides are infused. We used CL recording to demonstrate in rat heart that: (i) different diets may lead to different heart sensitivity to an oxidative stress; and (ii) post‐ischaemic reoxygenation induces an oxidative stress. CL emission induced by an oxidative stress is accompanied by an increased release of eicosanoids. However, while non‐steroid anti‐inflammatory drugs (aspirin, indomethacin and ibuprofen) prevented eicosanoid release, these compounds dramatically enhanced hydroperoxide‐dependent CL. The nature of this phenomenon is still obscure, but the increase of steady‐state concentration of excited species caused by anti‐inflammatory drugs seems to be pathophysiologically relevant, since in all our experimental conditions tissue damage was proportional to CL emission rate.
Localization of the luminol‐dependent chemiluminescence reaction in human granulocytes
Authors: Dahlgren Claes , Follin Per , Johansson Agneta , Lock Ronny , Orselius Kristina 
Published: May 2005

AbstractThe granulocyte luminol‐dependent chemiluminescence (CL) reaction is linked to the enzyme myeloperoxidase reacting with products of the respiratory burst activation. The results presented in this paper, show that the light generated in granulocytes originate both from intracellular and extracellular reactions; however, depending on the stimulus used the one or the other will dominate the activity measured. Furthermore, lysosomal fusion is proposed to be required for the intracellular CL reaction.
A rapid chemiluminescent DNA hybridization assay for the detection of Chlamydia trachomatis
Authors: Clyne J. M. , Running J. A. , Stempien M. , Stephens R. S. , Akhavan‐Tafti H. , Schaap A. P. , Urdea M. S. 
Published: May 2005

AbstractWith an estimated 3–4 million new cases per year, human infections from Chlamydia trachomatis are probably the most prevalent sexually transmitted disease (STD) in the United States. Diagnosis of Chlamydia is usually conducted by tissue culture methods. Direct immunofluorescence and ELISA tests have become available, but there remains a need for a test with better specificity and sensitivity. In response to this need, we have developed a rapid DNA hybridization assay using synthetic oligonucleotide probes to detect the presence of the Chlamydia trachomatis specific 7.4 kb plasmid. The assay involves solution phase hybridization of unlabelled probes, rapid capture of the probe‐target duplex onto a microtitre dish surface, a new signal amplification technique that employs chemically cross‐linked oligonucleotides, and an alkaline phosphatase labelled probe. Signal is obtained by reacting the labelled probe‐target complex with an enzyme triggerable dioxetane substrate. Detection of the chemiluminescent output is performed either with a luminometer or by exposure to instant film. All 15 serovars of Chlamydia trachomatis react positively while organisms known to co‐inhabit the human urogenital tract react negatively.
A review of bioluminescent ATP techniques in rapid microbiology
Authors: Stanley Philip E. 
Published: May 2005

AbstractUse of firefly luciferase to assay adenosine triphosphate (ATP) extracted from microorganisms provides an easy means to enumerate microbes within minutes. The small amount of light produced is proportional to ATP and thus microbial number.The average bacterium contains around 10−15 g ATP per cell. Present reagents permit detection of 103 cells per tube. Luminometers currently on the market detect about 10−12 g ATP.Proper extraction of ATP from the microbes is an essential part of any protocol, as is the removal of non‐microbial ATP from, for example, somatic cells also present in samples. The technique may be applied to a wide range of samples, for example food and beverages and clinical samples such as urine.The ATP assay gives a global measure of microbial numbers, i.e. it is not species specific unless a species separation step is included in the protocol.
Coupled reactions for the determination of analytes and enzymes based on the use of luminescence
Authors: Roda A. , Girotti S. , Ghini S. , Carrea G. 
Published: May 2005

AbstractImmobilized enzymes are widely used in the clinical laboratory in the assay of several analytes and enzymes. The use of immobilized enzymes makes these reagents recoverable and re‐usable, and in most cases increases their stability and catalytic activity. In conjunction with bioluminescent enzymes (firefly and bacterial luciferases) and chemiluminescent catalyst (peroxidase) we set up high‐sensitive flow methods based on the use of nylon tube coil or epoxy methacrylate column as solid support.All the NAD(P)/NAD(P)H‐dependent dehydrogenases (bacterial luciferase), ATP‐dependent enzymes (firefly luciferase) and oxidases producing H2O2 (peroxidase) can be immobilized and a large variety of analytes have been sensitively measured. As an alternative format we also reported a dry chemistry method in which all the enzymes, substrates and cofactors are ready to use, supported on dry cellulose disks. Methodological problems such as flow conditions, stability, pH, ionic strength and analytical performances are also reported.
Detection of chemiluminescence in lipid peroxidation of biological systems and its application to HPLC
Authors: Miyazawa Teruo , Saeki Rie , Inaba Humio 
Published: May 2005

AbstractA combined system of chemiluminescence detection and high performance liquid chromatography (CL–HPLC) was developed to determine primary peroxidation products in biological tissues, such as phosphatidylcholine hydroperoxide (PCOOH). The CL–HPLC assay consists of separation of lipid classes with HPLC and detection of hydroperoxide‐specific chemiluminescence. Hydroperoxides react with heme compounds to produce oxidants as suggested by our early studies on tissue low‐level chemiluminescence in which singlet molecular oxygen is generated as one of the excited species in several biological systems involving free radical events. In the CL–HPLC method, a cytochrome c–luminol mixture was used as a hydroperoxide‐specific luminescent reagent, and the quantification of hydroperoxide was performed by detecting chemiluminescence due to the luminol oxidation caused by the oxidant produced during the lipid hydroperoxides with heme. The detection limit of PCOOH was 10 pmole hydroperoxide–O2. PCOOH in normal human blood was found to be 10–500 pmol/ml plasma and significantly higher levels of PCOOH were observed in some hospitalized patients.
The use of a CCD imaging luminometer in the quantitation of luminogenic immunoassays
Authors: Leaback D. H. , Haggart R. 
Published: May 2005

AbstractWe describe the application of a CCD Imaging Luminometer (CCDIL) for the detection and quantitation of rapid, simultaneous, peroxidase‐linked chemiluminescent immunoassays of multiple samples of human serum alphafetoprotein (AFP). Results from some analogous immunoassays (total IgE, and thyroid stimulating hormone (TSH)) are included for comparison.Values for precision and antigen concentration obtained using the CCDIL and colorimetric versions of the immunoassays, on human serum samples, were in good agreement.The flexibility of the CCDIL is demonstrated; its ability to detect and quantitate antigen (particularly AFP) on a variety of solid phases is indicated.The work on the AFP immunoassays illustrates not only the flexibility of the CCDIL for sample presentation on a variety of solid phase systems, but also some relative merits of such systems.
Bioluminescence in oceanology
Authors: Gitelson I. I. , Levin L. A. 
Published: May 2005

AbstractFor analytical purposes bioluminescence can be used in three main ways: 1. luminescence measurement of bioluminescent system components isolated in vitro; 2. determination of luminous organisms' reaction to the in vivo test‐action; 3. measurement of bioluminescence in marine ecological systems.The majority of the reports of this Symposium are dealing with the first two topics. The aim of our presentation is to draw attention to the third one. The possibilities of bioluminescent analysis are wider than its traditional scheme of applications in the laboratory, when the emitting system is withdrawn from a native source and is placed in a cuvette of the light measuring device. The reverse scheme is also possible, i.e. the device can be introduced into light emitting system such as a marine biocenosis–the community of the sea inhabitants–where we obtain a highly sensitive and rapid means of gaining the information on the vital activity of marine ecosystems, i.e. their spatial structure, rhythms, man's influence upon them, etc. The present communication will consider the possibilities of this form of bioluminescent analysis.
Methodological problems of direct bioluminescent ATP assay in platelets and erythrocytes
Authors: Girotti Stefano , Ferri Elida , Cascione Maria Loredana , Comuzio Sergio , Mazzuca Augusto , Orlandini Alessandro , Breccia Alberto 
Published: May 2005

AbstractDirect bioluminescent ATP determination in platelets and erythrocytes involves the study of different parameters which are discussed here. Some parameters are linked to the bioluminescent reaction and to the analyte (ATP); others have regard to the biological matrix. The composition of bioluminescent reagents and the preparation and conservation of the ATP standard, also in the presence of excipients, are among the first given. Matrix problems involve cell characteristics related to age and form, lysis resistance and the possible formation of aggregates (platelets) that may inhibit the complete release of ATP. For these reasons we used the most efficient ATP release agent with the lowest inhibitory effect on luciferase. The data obtained correlate well with a bioluminescent method requiring extraction with ethanol/EDTA, and therefore more time, for ATP determination in platelets and erythrocytes.
Unusual luminescent properties of odd‐ and even‐substituted naphthyl‐derivatized dioxetanes
Authors: Edwards B. , Sparks A. , Voyta J. C. , Bronstein I. 
Published: May 2005

AbstractWith the advent of enzymatically induced chemiluminescence and improved instrumentation for luminometry, ultrasensitive detection of a wide variety of analytes is now possible using standard immunoassay and DNA probe formats. Model molecular orbital calculations and literature precedent suggest that the singlet efficiencies observed upon decomposition of dioxetanes appended with donor substituted aromatic moieties are dependent on substitution pattern. We have recently discovered, in a series of 3‐(2′‐spiroadamantane)‐4‐methoxy‐4‐acetoxynaphth‐2′‐yl‐1,2‐dioxetanes, that enzymatic generation of a nonconjugated, charge transfer excited state results in luminescence of markedly different properties than that observed from an isomeric, conjugated excited state. An example of the former type, 3‐(2′‐spiroadamantane)‐4‐methoxy‐4‐(7″‐acetoxy)naphth‐2′‐yl‐1,2‐dioxetane (1) emitting at 550 nm, not only provides an increase in ΦCL but exhibits a dramatic bathochromic shift of 80–110 nm from the 460 nm emission of the conjugated isomer 3‐(2′‐spiroadamantane)‐4‐methoxy‐4‐(6″‐acetoxy)naphth‐2′‐yl‐1,2‐dioxetane (2).These developments, along with the attendant glow‐type luminescence kinetics displayed during the enzymatic decomposition of the new ‘odd‐pattern’ dioxetane, allow the design of simple protocols capable of simultaneous or ‘multichannel’ detection of several analytes.
Fusion of luxA and luxB and its expression in E. coli, S. cerevisiae and D. melanogaster
Authors: Almashanu S. , Musafia B. , Hadar R. , Suissa M. , Kuhn J. 
Published: May 2005

AbstractLuciferase from Vibrio harveyi is encoded by two adjacent genes, luxA and luxB. The two genes were fused by replacing a segment extending from near the end of luxA into the N‐terminal end of luxB by a synthetic oligonucleotide. The construction removed the TAA stop codon at the end of luxA, the intervening region of 26 base pairs, and the initial methionine of luxB. A Smal site was included at the junction between the two genes and an Aatll site was created near the end of luxA without altering its amino acid sequence. In Escherichia coli the fused luxAB gene could be expressed to produce functional luciferase that gave about 20% of the activity in cells without the fusion.An out‐of frame ATG exists close to and preceding the ATG of the luxA gene. This was removed and the entire fused gene bracketed by several restriction enzyme sites.The fused luxAB gene was successfully expressed in Saccharomyces cerevisiae and Drosophila melanogaster by transferring it to appropriate plasmid vectors.
The effect of detergents on firefly luciferase reactions
Authors: Simpson W. J. , Hammond J. R. M. 
Published: May 2005

AbstractThe reaction rate of ATP‐limited firefly luciferase‐catalysed reactions, is affected by the presence of detergents. Anionic detergents inhibit luciferase activity without causing significant enzyme inactivation during the reaction. Cationic detergents increase reaction rate several‐fold with a sharply defined optimum concentration of detergent for the effect. However, cationic detergents inactivate firefly luciferase during the reaction, resulting in a continuously decreasing reaction rate. Under such conditions, peak light intensity must be used as an indication of initial reaction rate. The inactivation rate increases with increasing detergent concentration. Non‐ionic and zwitterionic detergents increase reaction rate over a broad range of detergent concentrations. Enzyme stability during the reaction is not affected by non‐ionic detergents and only affected by zwitterionic detergents at high detergent concentration. Cyclodextrins, which can increase reaction rates of some chemiluminescent reactions, have little effect on firefly luciferase activity.Assays for ATP using firefly luciferase must be internally standardized by the constant addition technique in which a known amount of ATP is added to the test sample, since external calibration of such assays, by reference to a previously prepared standard curve, can lead to imprecision when detergents are present.
Chemiluminescent 10‐methyl‐acridinium‐9‐(N‐sulphonylcarboxamide) salts. Synthesis and kinetics of light emission
Authors: Mattingly Phillip G. 
Published: May 2005

Abstract10‐Methyl‐acridinium‐9‐(N‐sulphonylcarboxamide) salts are prepared via acylation of sulphonamides with acridine‐9‐carboxylic acid chloride and subsequent N‐alkylation with methyl triflate. Substituents on the sulphonamide component were varied to show the effect of steric and electronic factors on the kinetics of light output. The lifetime of the chemiluminescence light output ranged from 1 to 50 seconds for the 15 compounds reported. The long‐term stability of the new compounds was superior to the phenyl ester counterparts.