1. OBJECTIVES/METHODS: Bleed vaccinees annually; test serum for rabies- neutralizing antibody by the Rapid Fluorescent Focus Inhibition Test (RFFIT); boost those with < 1.0 International Unit rabies-neutralizing antibody/ml serum with HDCS Rabies Vaccine (Institut Merieux). In FY91, this project was partially suspended due to loss of personnel and training required for their replacement. During this time, NIH sent their sera to the Rabies Laboratory at Kansas State University, Manhattan, Kansas. We resumed testing these sera late this year so the numbers are lower than usual. 2. RESULTS: Distribution of human serum quantified for rabies-neutralizing antibody in IU/ml: Employing Agency (serum source) Number RFFIT Tests CBER-FDA 30 NIH 24 TOTAL HUMAN SERUM TESTS 150 GRAMT=Z01BE01003 An ELISA has been developed to measure the amount of anti-tetanus antibodies in serum from guinea pigs immunized with adsorbed biological products containing the tetanus toxoid component. The purpose of this portion of the study is to determine variation among plates and to examine the reproducibility of each relative potency value. The current ELISA standard is guinea pig serum diluted to about 6 units per ml. The unknowns will consist of the standard and the standard diluted to 1,2,3,4, and 5 units. Each sample will be assigned a code. This code identifies the plate where the unknown is located. The standard and the unknowns will have a preliminary screening (rough test). All samples will be tested at four dilutions (1:500; 1:1000; 1:5000; and 1:10,000) in duplicate. The preliminary testing of the standard or an unknown sample should provide adequate information to determine the starting dilution for the fine test. From these starting dilutions, a fine test will be performed to generate O.D. values for each point using a 1:1.5 dilution scheme. The plate is read at 410 nm and analyzed by a computer program (the parallel line assay) designed specifically for this application. The program assigns a unit value for the unknowns based on the standard and gives the 95% confidence limits for the value of the unknowns. The unknowns are decoded and compared to the relative potency values generated by the computer program. The variation among plates and the reproducibility of all replicates are analyzed statistically and interpreted. Automated vs. manual dilutions are being compared for accuracy and precision.