Salgado J, Gilabert A, Castell M, Castellote C, Queralt J
Department de Ciències Fisiològiques Humanes i de la Nutrició, Universitat de Barcelona, Spain.
Allergol Immunopathol (Madr). 1988 Mar-Apr;16(2):95-8.
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.
已开发出两种间接微量酶联免疫吸附测定（microELISA）方法来测定大鼠血清中卵清蛋白（OA）特异性IgG和总OA特异性抗体。动物经皮下注射用明矾悬浮的OA免疫，并腹腔注射百日咳博德特氏菌（Bp）。该研究使用免疫后14天采集的血清样本进行。测量了抗OA IgG、抗OA Ig（通过间接ELISA法）、总IgE和总IgG（通过ELISA夹心法）以及抗OA特异性IgE（通过被动皮肤过敏试验）。包被固相的OA浓度为1.25微克/毫升。所开发的间接ELISA法测量OA特异性抗体的最佳血清稀释度在1/640 - 1/1280之间，两种方法均如此。所用结合物浓度分别为1/12800（过氧化物酶 - 抗大鼠IgG）和1/600（过氧化物酶 - 抗大鼠Ig）。综合两种方法，批内和批间变异系数在1.98%至6.75%之间波动。所采用的免疫程序使抗OA Ig、抗OA IgG和特异性IgE水平显著升高。免疫未改变总IgG水平，而总IgE值显著更高。这种升高主要可归因于Bp诱导的非特异性IgE产生。本研究中提出的微量ELISA方法是测量OA特异性抗体的放射性标记技术的合理替代方法。
