EVALUATION OF ANTIGENS IN THE ELISA INHIBITION ASSAY FOR NATURAL RUBBER LATEX PROTEINS
V. J. Tomazic-Jezic, A. D. Lucas, FDA/CDRH, Rockville
In the process of development of a ELISA Inhibition assay for the quantitation of the natural rubber latex (NRL) proteins, we investigated properties of proteins from ammoniated (AL) and non-ammoniated (NAL) raw latex, as coating antigens, immunizing antigens and reference antigens. The anti-NRL sera were produced by immunizing rabbits with AL, NAL and a mix of both antigens. Determination of affinity of rabbit antisera for binding to these antigens indicated strong preference of both anti-AL and anti-NAL antibodies to bind the immunizing antigen, while anti-Mix antiserum bound similarly to both antigens. Evaluation of the three sera in the ELISA inhibition assay with AL and NAL proteins as coating and reference antigens, further confirmed this finding. Anti-AL serum demonstrated a strong preference in inhibiting and binding to AL antigen, as compared to either NAL antigen or mix. In the case where NAL was a coating antigen, standard curves for both inhibition antigens (AL and NAL) with all three sera were more linear and uniform. Next we investigated the capacity of different anti-sera to inhibit proteins in glove extracts, comparing two coating antigens, AL and NAL. We observed a significant difference in the appearance of the inhibition curves of glove extracts with these two coat antigens. When NAL was used as a coat antigen, the inhibition curves by all three sera were similar. The results indicate that the selection of an appropriate combination of anti-serum, inhibiting and coating antigen is critical for the accurate measurements of the NRL proteins in the finished products.