Journal of Immunological Methods, 60 (1983) 213-220 213 Elsevier Biomedical Press Measurement of Parasite-Specific Antibody Responses using a Tritiated Avidin-Solid Phase Radioimmunoassay R.L. Tarleton 1 and R.E. Kuhn Department of Biology, Wake Forest University, Winston -Salem, NC 27109, U.S.A. (Received 13 October 1982, accepted 19 December 1982) A solid phase radioimmunoassay (RIA) to measure parasite-specific antibody responses has been developed. The assay utilizes 3H-labeled avidin as the indicator molecule and represents a marked improvement in detection of specific antibodies to Trypanosoma cruzi over previously used RIA. Avidin is easily labeled to high efficiency (3.6 × 103 cpm/#g) under very mild conditions using N-succinimidyl-[2,3- 3H]propionate, and the high efficiency of avidin-biotin binding coupled with the increased" safety and longer half-life of the labeled reagent give this method advantage over currently used 125I-labeled Ab RIAs. Key words: radioimmunoassay -- tritium -- avidin- biotin -- parasite -- Trypanosoma cruzi Introduction Quantitation of antibodies specific for parasites in infected hosts has most recently been accomplished using primarily either enzyme-linked immunoadsorbent assay (ELISA (Hamilton et al., 1981; Sethi and Brandis, 1981)) or radioimmunoas- say (RIA (Kim et al., 1980; Hamilton et al., 1981; Sethi and Brandis, 1981)) techniques. Our initial attempts to determine the level of parasite-specific antibody in the serum of infected animals using 125I-RIA was hampered by the high background levels of binding of immunoglobulins in normal serum controls. With the combined use of tritium labeling and the avidin-biotin system, we have developed a highly sensitive, relatively safe and easy assay for determining the level of parasite-specific antibody in hosts infected with the protozoan Trypanosoma cruzi. i Supported in part by a National Science Foundation Graduate Fellowship. 0022-1759/83/0000-0000/$03.00 © 1983 Elsevier Science Publishers