A new commercial glycoprotein G-based enzyme immunoassay (gG-EIA) was compared with Western blotting (WE) for detection of herpes simplex virus type 1 (HSV-1) or HSV-2 type specific antibodies in 193 serum samples. Sensitivity for HSV-1 was 95%; specificity was 96%. Sensitivity for HSV-2 was 98%; specificity was 97%. Twelve of 13 serum samples with equivocal gG-EIA results were serotyped by WB.