The enzyme-linked immunosorbent assay (ELISA): A measure of antibody concentration or affinity?

Butler, John E.; Feldbush, Thomas L.; McGivern, P.L.; Stewart, Nancy

doi:10.1016/0161-5890(78)90053-6

Cited by 185 publications

(56 citation statements)

References 11 publications

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“…The enzymatic reaction was developed with an ABTS (Boehringer-Mannheim)-based chromogen and measured by a computer-controlled photometer (Multiscan MCC-340; Flow Lab, Inc., McLean, Va.) (12). Antibody concentrations were expressed as ELISA units (EU) using the positive reference standard method (6).…”

Section: Methodsmentioning

confidence: 99%

Immunodiagnostic Identification of Dairy Cows Infected withPrototheca zopfiiat Various Clinical Stages and Discrimination between Infected and Uninfected Cows

2001

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Protothecosis is a severe form of mastitis in cattle that is caused by colorless algae of the genus Prototheca. So far, no suitable serological test for the identification of infected animals is available for routine diagnosis. In this study an indirect enzyme-linked immunosorbent assay (ELISA) for the identification of infected cows and for discriminating among infected cows at various clinical stages was developed. Immunoglobulin G (IgG) in serum and IgA and IgG1 in whey were used as antibody isotypes. The ELISA was evaluated using serum and whey from animals at different clinical stages of infection. A total of 12 cows with acute clinical manifestation of protothecal mastitis, 22 cows with clinical signs of chronic mastitis, 40 Prototheca zopfii-negative cows, and 18 cows with chronic clinical signs and earlier cultures positive for P. zopfii but with presently negative culturing results were investigated. A sensitivity of 96% and a specificity of 94% were calculated for the ELISA based on IgA levels. Intra-assay and interassay variations were calculated to be 6.08 and 6.32%, respectively. Based on these data, this ELISA was found to be suitable for discrimination between infected and uninfected animals and might therefore be useful for screening affected herds.

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Section: Methodsmentioning

confidence: 99%

Immunodiagnostic Identification of Dairy Cows Infected withPrototheca zopfiiat Various Clinical Stages and Discrimination between Infected and Uninfected Cows

2001

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“…In the present study, bovine RSV-specific antibody titres were determined by ELISA using the end-point titration method. The end point titration method has generally been preferred to the single dilution method because it is probably less influenced by antibody affinity and antibody titre [35][36][37].…”

Section: Lung Lavagementioning

confidence: 99%

Class-specific antibodies to bovine respiratory syncytial virus in experimentally infected lambs

Sharma

Woldehiwet²

1992

Epidemiol. Infect.

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SUMMARYEnzyme-linked immunoabsorbent assay (ELISA) was used to titrate virusspecific IgG, IgM and IgA levels in nasal secretions, lung lavage fluids and serum samples sequentially obtained from lambs experimentally infected with bovine respiratory syncytial virus (RSV). Virus-specific IgG and IgM responses were measured by the indirect double antibody sandwich ELISA using anti-bovine RSV monoclonal antibody, as capture antibody, and peroxidase-conjugated antisheep IgG and anti-sheep IgM. Virus-specific IgA antibodies were measured by antibody capture assay using anti-sheep IgA (a-chain specific) and anti-bovine RSV monoclonal antibodies.

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“…It has been observed that standard ELISAs measure high avidity antibodies whereas amplified ELISAs can detect low avidity antibodies.24 In our study the prolonged incubation of serum for 48 hours with the poly-L-lysine method, and for 24 hours with the mHSA method, suggests that low avidity antibodies were being detected. It was possible to reduce this serum incubation time to one hour in the poly-L-lysine method, and obtain a similar absorbance value, in the presence of 10% polyethylene glycol.…”

Section: Reproducibiitiymentioning

confidence: 74%

Two enzyme linked immunosorbent assays for detecting antibodies against meningococcal capsular polysaccharides A and C.

Akinwolere

Kumararatne

Bartlett

et al. 1994

Journal of Clinical Pathology

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Aims-To evaluate two of the recent methods of coating microtitre plates in the enzyme linked immunosorbent assay (ELISA) for detecting human antibodies against meningococcal capsular polysaccharides A and C with a view to validating a specific meningococcal antibody assay for routine clinical use. Methods-Two four-layer ELISA protocols were standardised: one method utilised meningococcal polysaccharides conjugated to poly-L-lysine polypeptide for coating the microtitre plates; another used polysaccharides mixed with methylated human serum albumin (mHSA). Titration curves were plotted for the ELISAs and the squared Pearson correlation coefficient (R2) was used to determine the degree of accuracy of fit of the curves. Specificity tests were performed by inhibition and adsorption studies. Results-Both methods gave good titration curves with a high R2 of >0-98, indicating a high degree of accuracy in forming the curves. The titration end point after vaccination, obtained by the mHSA method, was 20 times higher, however, than that obtained by the poly-L-lysine method. Specificity tests showed that in the ELISA using polysaccharide/poly-L-lysine, antibody activity of a pre-vaccination serum sample was inhibited by 37%, and of post-vaccination serum by 50%/ with 1000-fold excess antigen. Antibody activity (postvaccination) was reduced by 51% and 59o/o, respectively, by adsorption with antigen-coated Sepharose beads or adsorption with suspensions of killed meningococci. In contrast, antibody activity of a pre-vaccination serum was inhibited by 60% and a post-vaccination serum by 90% in ELISA employing polysaccharides mixed with mHSA. Reproducibility was better with the use of methylated human serum albumin than with poly-L-lysine; the former showed intrabatch and interbatch coefficients of variation of 4% and 2%, respectively, compared with 43% (intrabatch) and 16% (interbatch) obtained with the poly-Llysine. Conclusion-It is concluded that the antibody assay using meningococcal polysaccharides groups A and C mixed with mHSA is much better than that using polysaccharides coupled with poly-Llysine.

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The enzyme-linked immunosorbent assay (ELISA): A measure of antibody concentration or affinity?

Cited by 185 publications

References 11 publications

Immunodiagnostic Identification of Dairy Cows Infected withPrototheca zopfiiat Various Clinical Stages and Discrimination between Infected and Uninfected Cows

Immunodiagnostic Identification of Dairy Cows Infected withPrototheca zopfiiat Various Clinical Stages and Discrimination between Infected and Uninfected Cows

Class-specific antibodies to bovine respiratory syncytial virus in experimentally infected lambs

Two enzyme linked immunosorbent assays for detecting antibodies against meningococcal capsular polysaccharides A and C.

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