Quantitation of antigen-specific immunoglobulin G in human serum: II. Comparison of radioimmunoprecipitation and solid-phase radioimmunoassay techniques for measurement of immunoglobulin G specific for a complex allergen mixture (yellow jacket venom)

Abstract

In this report, the radioimmunoprecipitation (RIP) assay and the Staph A (protein A from Staphylococcus aureus) solid-phase radioimmunoassay (SPRIA) are compared for their ability to measure IgG specific for a complex antigen mixture. Sera from 32 yellow jacket venom (YJ)-allergic patients were evaluated for IgG anti-YJ using both radioassay methods. Twenty-six of the 32 sera gave concordant results in both assay systems. Four of the six discrepant sera were 5- to 12-fold higher by RIP analysis while the remaining two sera were 2 and 4.3 times higher in the SPRIA. Serum-dilution curves of these six discrepant sera in the RIP assay demonstrated nonparallelism and differential plateau binding ranging from 40% to 140% of the reference serum plateau level. In contrast, the same six discordant sera evaluated in the SPRIA produced parallel serum-dilution curves. To assess the validity of each method, absorption experiments and Staph A solid-phase elution technique (SPET) were performed on each discrepant serum. Absorption experiments with insolubilized YJ antigen showed that all IgG anti-YJ detectable in the RIP was removed. Furthermore, results from the Staph A elution technique demonstrated the same rank order and relative potency of the six discrepant sera as shown by the SPRIA but not by the RIP analyses. These findings suggest that for some YJ sera, the SPRIA gives more accurate estimates of IgG antibody content than does the RIP assay. In addition, the SPRIA presents several advantages over the RIP in the measurement of specific IgG in complex antigen mixtures: (1) The problems of radioiodination reproducibility and low immunoreactivity that are commonly seen with crude antigenic mixtures are avoided. (2) Antigen-excess conditions are employed, thereby assuring sufficient antigen to quantify all antibody specificities. (3) IgE and IgG measurements in the same sera are more comparable since the same antigen-sorbent can be used in both the radioallergorsorbent test (RAST) for IgE and Staph A SPRIA for IgG. (4) Only one radiolabeled detection protein is required for evaluation of many antigen systems, thus alleviating the need for frequent radioiodination of multiple complex antigen mixtures. (5) Problems of nonparallelism and differential plateau binding that were observed in a minority of sera in the RIP assay for YJ antibody may be circumvented. We conclude, therefore, that the SPRIA has distinct advantages that may commend its use over RIP in the assessment of specific IgG in some complex antigen systems.