Better screening tests for syphilis are required to help ensure safer blood supply. Reverse screening for syphilis using specific test like chemiluminescence immunoassay (CLIA) allows automation for high sample load testing, followed by testing of positive sera with a non-treponemal. In case of discordant result between the two tests, the sample is tested with another confirmatory specific test. The objective of this study was to evaluate the performance of automated Vitros Syphilis Treponema pallidum antibody chemiluminescence immunoassay as a screening test in blood donors using reverse algorithm for testing syphilis. A total of 16362 blood donors were screened for syphilis by CLIA. The CLIA positive samples and 100 CLIA negative samples were further tested by Rapid Plasma Reagin Test (RPR) and Treponema pallidum haemagglutination test (TPHA). 155 (0.95%) sera were positive by CLIA among which 34(21.93%) were reactive by RPR and 32 (20.6%) were confirmed positive by TPHA. 121(78.06%) samples yielded discordant result with RPR, of which 10 (6.45%) were confirmed positive by TPHA. The sensitivity of CLIA in comparison to TPHA was 100%, specificity 46.95%, positive predictive value 27.1%, negative predictive value 100% and kappa coefficient 0.2257 (poor agreement). CLIA detected 10 (6.45%) cases of syphilis which would have been missed if RPR was used as the initial screening test. CLIA can be employed for screening syphilis in blood donors for its good sensitivity, ease of automation and short turn-around-time.