A flow microsphere inhibition immunoassay (FMII) for detecting paratope binding anti-idiotypic antibodies.
A simple, rapid and reproducible flow microsphere inhibition immunoassay (FMII) has been developed to detect the ability of paratope specific anti-idiotypic antibody (anti-Id or AB2) to inhibit antigen binding to the corresponding paratope of the Id (AB1). To evaluate the FMII as a measurement of paratope binding anti-Id, an avian model was used to produce Id and anti-Id antibodies for the study. Both antibody to bovine serum albumin (BSA Id) and anti-BSA Id were produced in white leghorn chickens and affinity isolated from egg yolks. The anti-BSA Id samples were incubated with BSA Id coated microspheres, then without rinsing, fluoresceinated BSA (BSA-FITC) was added for a short incubation period and the resulting decrease in fluorescent intensity was used to calculate the extent of inhibition. For validation, statistical comparisons of the line equations generated by BSA dilution curves and anti-BSA Id dilution curves were performed. Replications within each ligand were not significantly different which indicated the assay was reproducible for determining the presence of paratope reactive anti-BSA Id used in this model.
Brown
J E
JE
College of Veterinary Medicine, Mississippi State University, MS 39762.
Ainsworth
A J
AJ
D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000032:analysis*; D000918:Antibody Specificity; D001666:Binding Sites, Antibody; D002417:Cattle; D002645:Chickens; D005069:Evaluation Studies as Topic; D007118:Immunoassay / Q000379:methods*; D007130:Immunoglobulin Idiotypes; D008863:Microspheres; D012710:Serum Albumin, Bovine / Q000276:immunology
D000888:Antibodies, Anti-Idiotypic; D007130:Immunoglobulin Idiotypes; D012710:Serum Albumin, Bovine