A urease-based enzyme-linked immunosorbent assay (ELISA) has been developed for the detection of reagin antibodies in serum. Visuwell Reagin (ADI Diagnostics Inc., Rexdale, Ontario, Canada) is a non-treponemal screening test for the serodiagnosis of syphilis which has the benefits of large batch testing, automatability, and objective interpretation of results. Unheated, undiluted sera are incubated in 96-well microtiter plates coated with a modified cardiolipin-lecithin-cholesterol antigen. Antibody bound to the plate is detected by an anti-human immunoglobulin G-urease conjugate. The procedure consists of three steps, with a total test time of 60 min. Visuwell Reagin ELISA was compared with the Venereal Disease Research Laboratory (VDRL) test and the reagin screening test (RST) with the following results. For ELISA versus the VDRL test, the sensitivities for untreated syphilis (n = 37) were 97.3% for both ELISA and the VDRL test, the confirmatory positive values (n = 79) were 84.8% for ELISA and 72.2% for the VDRL test, and the specificities for normal samples (n = 1,327) were 98.8% for ELISA and 99.5% for the VDRL test. For ELISA versus RST, the sensitivities for untreated syphilis (n = 57) were 94.7% for ELISA and 87.7% for RST, the confirmatory positive values (n = 26) were 96.2% for ELISA and 92.3% for RST, and the specificities for normal samples (n = 1,891) were 99.6% for ELISA and 99.3% for RST. The overall concordance values of ELISA with VDRL test and RST were 96.7 and 97.9%, respectively. The specificity of ELISA compared with that of RST may be underestimated, since confirmatory data were not available for all apparent false-positive samples. Visuwell Reagin had increased sensitivity and similar specificity compared with flocculation tests.