An investigation of the use of urease-antibody conjugates in enzyme immunoassays

Chandler, Howard M.; Cox, John C.; Healey, K.; MacGregor, A.; Premier, Robert; Hurrell, John G.R.

doi:10.1016/0022-1759(82)90140-5

Cited by 103 publications

(32 citation statements)

References 9 publications

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“…The urease system [Chandler et al, 1982] was used in all enzyme-linked immunosorbent assays. Reac tions were graded visually as either negative (bright yellow), posi tive (purple), or strong positive (dark purple).…”

Section: Development O F Monoclonal Antibodiesmentioning

confidence: 99%

Development of Monoclonal Mouse Antibodies Specific for Allergenic Components in Ryegrass <i>(Lolium perenne)</i> Pollen

Smart

Heddle

Zola

et al. 1983

Int Arch Allergy Immunol

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The development of monoclonal mouse antibodies against ryegrass (Lolium perenne) pollen allergens is described. Hybridoma colonies secreting antibodies specific for allergenic components were detected using an enzyme-linked immunosorbent assay. Positive colonies were cloned and expanded. The pollen components with which the monoclonal antibodies interact were identified and characterised following sodium dodecyl sulphate polyacrylamide gel electrophoresis and electrophoretic transfer to nitrocellulose. In this paper six monoclonal mouse antibodies are described. Three antibodies interact with a single molecule of between 30,000 and 35,000 daltons. One antibody interacts with a component of 16,000 daltons whereas the remaining two antibodies react with more than one component, one reacting with two components at 28,000 and 30,000, and the other with five components having molecular weights between 18,000 and 71,000 daltons.

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Section: Development O F Monoclonal Antibodiesmentioning

confidence: 99%

Development of Monoclonal Mouse Antibodies Specific for Allergenic Components in Ryegrass <i>(Lolium perenne)</i> Pollen

Smart

Heddle

Zola

et al. 1983

Int Arch Allergy Immunol

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“…The monolayers were then washed twice each with PBS-Tween, 0.85 % saline, and distilled water. Following addition of the urease substrate (0.008 % bromocresol purple, 0.15 mM NaOH, 16.0 mM urea, 0.2 mM ethylenediaminetetraacetate (EDTA; Chandler et al 1982), the monolayers were incubated at 37 °C and OD570 readings were taken at 0.5, 1.0, and 1.5 h. Wells with a change in OD greater than 0.1 OD units above the control value were considered positive, based upon OD values obtained from a panel of cell lines previously tested by flow k p cytometry (Fig. 3).…”

Section: Monoclonal Antibodies (Mab)mentioning

confidence: 99%

Structure function analysis of theH-2 Ab p gene

et al. 1991

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The gene encoding the H-2 Ap class II beta chain was isolated from a B10.P genomic library and sequenced. This gene was also used to construct transfectants of the CH12 lymphoma clone CH12.LX, which express the Abp gene product in association with the endogenous A alpha k chain. We present here the first report of the complete nucleotide coding sequence of Abp. The predicted amino acid sequence of Abp reveals only five residues different from Abq, four of which are present in the mature peptide. These four amino acid changes could account for the differential susceptibility of H-2q vs H-2p mice to the development of collagen-induced arthritis (CIA). Antibodies specific for the transfected Abp protein induce CH12.LX cells to secrete immunoglobulin in the presence of antigen. Comparison of the amino acid sequence with other A beta chains that have been tested in signal transduction experiments suggests that amino acid 9 may be important to the signaling ability of class II A molecules.

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“…Urease is widely used for determination of urea by bioanalytical methods and in immunoassays as an enzyme label for many antigens [2,3]. Despite the great importance of urease in biotechnology and medicine, the detailed mechanism of urea hydrolysis by urease is still not clarified, and the structure of the metal site, containing two nickel atoms, is not precisely known [1,4,5].…”

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confidence: 99%

Inhibition of soybean urease by polycarbonyl compounds

2005

Appl Biochem Microbiol

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Soybean urease (EC 3.5.1.5) is a homohexameric enzyme (containing six subunits with molecular weights of 90770 Da and two nickel atoms each), which catalyzes urea hydrolysis by water, resulting in the formation of ammonium cations and carbonate anions [1]. Urease is widely used for determination of urea by bioanalytical methods and in immunoassays as an enzyme label for many antigens [2,3]. Despite the great importance of urease in biotechnology and medicine, the detailed mechanism of urea hydrolysis by urease is still not clarified, and the structure of the metal site, containing two nickel atoms, is not precisely known [1,4,5].An important tool for the investigation of the catalytic mechanism of urease is inhibition and activation of this enzyme. The main groups of urease inhibitors, specificity and efficiency of urease activity were discussed in our earlier review [6]. It was found that urease was produced by many microorganisms and was associated with such pathologies as duodenal and gastric ulcers as well as a number of human and animal urinary diseases [5,7]. It was also demonstrated that there was a clear correlation between the inhibiting properties of the same compounds with respect to various ureases of microbial and plant origin. Therefore, the primary screening of potential inhibitors for urease can be performed more easily and quickly using catalytically active and thermally stable soybean urease [4,5,8].In earlier studies, we investigated the following urease inhibitors: polydisulfides of urea and thiourea, which compete with the substrate for binding to the urease active sites; amides of thiophosphoric acid, which were analogues of the substituted phosphorodiamidates (these compounds were the most effective urease inhibitors [4-6, 8]); cyclic β -triketones (CTKs); and fluoride anions. These inhibitors are organic and inorganic ligand chelators of nickel [9]. An important advantage of CTKs and fluoride anions is that they are nontoxic. It is obvious that the range of polycarbonyl compounds (PCCs), potential chelators of nickel ions, should be widened, and nickel ligands of polycarbonyl structure should be tested as urease inhibitors within a broad pH range, because the value of the inhibition constant for urease inhibition by fluoride anions significantly increases due to pH variation within the range from 3.85 to 6.45 [9]. This factor is particularly important for practical purposes because of the acidity of gastric medium in humans and animals.The practical aspect of this work is associated with selection of effective urease inhibitors for use as chemotherapeutical agents in gastroenterology and stopreagents in biochemical assay of urea (enzyme immunoassay and biosensors).The goal of this work was a comparative kinetic study of soybean urease inhibition by such organic chelators of nickel as polycarbonyl compounds, including oxalyldihydrazide (ODH), with two carbonyl groups, cyclic triketones I -VII with three carbonyl groups, and poly(disulfide-oxalyldihydrazide (poly(DSODH)) with 28 carbonyl gro...

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An investigation of the use of urease-antibody conjugates in enzyme immunoassays

Cited by 103 publications

References 9 publications

Development of Monoclonal Mouse Antibodies Specific for Allergenic Components in Ryegrass <i>(Lolium perenne)</i> Pollen

Development of Monoclonal Mouse Antibodies Specific for Allergenic Components in Ryegrass <i>(Lolium perenne)</i> Pollen

Structure function analysis of theH-2 Ab p gene

Inhibition of soybean urease by polycarbonyl compounds

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