ELISA for quantification of specific IgG and IgE antibodies to ovalbumin.
Two indirect microELISA methods have been developed to determine ovalbumin (OA)-specific IgG and total OA-specific antibodies in rat's serum. Animals were immunized with OA suspended in alum s.c. and administered i.p. with Bordetella pertussis (Bp). The study was performed using serum samples taken 14 days after immunization. Anti-OA IgG, anti-OA Ig (by indirect ELISA), total IgE and total IgG (by ELISA sandwich) and also anti-OA specific IgE (by passive cutaneous anaphylaxis test) were measured. The OA concentration to coat the solid phase was 1.25 micrograms/ml. The optimal serum dilution for measuring OA-specific antibodies by the indirect ELISAs developed was between 1/640-1/1280, for both methods. Conjugate concentrations used were 1/12800 (PO-anti rat IgG) and 1/600 (PO-anti rat Ig). Considering both methods, the intra and inter-assay variability coefficients oscillate between 1.98 and 6.75%. The immunization procedure used produces a marked increase in anti-OA Ig, anti-OA IgG and specific IgE levels. The immunization does not modify total IgG levels whereas total IgE values were significantly higher. This increase can be attributed chiefly to the non-specific IgE production induced by Bp. The microELISA methods proposed in this work, are reasonable alternatives radiolabelled techniques to measure OA-specific antibodies.
Salgado
J
J
Department de Ciències Fisiològiques Humanes i de la Nutrició, Universitat de Barcelona, Spain.
Gilabert
A
A
Castell
M
M
Castellote
C
C
Queralt
J
J
D000818:Animals; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D007114:Immunization; D007073:Immunoglobulin E / Q000032:analysis* / Q000276:immunology; D007074:Immunoglobulin G / Q000032:analysis* / Q000276:immunology; D008832:Microchemistry; D010047:Ovalbumin / Q000276:immunology*; D051381:Rats; D011919:Rats, Inbred Strains
D007074:Immunoglobulin G; D007073:Immunoglobulin E; D010047:Ovalbumin