To determine the accuracy of two currently used immunoassays and a newly developed optical immunoassay for rapid intrapartum detection of group B Streptococcal colonization compared with culture methods. Rayon-tipped swabs were used to collect specimens from the distal vagina of 502 women at admission for labor or rupture of membranes. Four tests were performed on specimens from the first 197 patients: culture in selective broth medium, semiquantitative culture on blood agar medium, and ICON Strep B and Quidel Group B Strep Test enzyme immunoassays. For the remaining 305 women, a fifth test, Strep B OIA, a newly developed optical immunoassay, was also performed. The prevalence of group B Streptococcal vaginal colonization was 25.1% by selective broth medium and 17.3% when swabs were plated directly onto blood agar medium, giving the latter method a sensitivity of 69%. When compared with selective broth medium Results , the sensitivities of the rapid immunoassays were 12% (Quidel), 15% (ICON), and 37% (Strep B OIA). These values rose to 16% (Quidel), 21% (ICON), and 53% (Strep B OIA) when compared with nonselective blood agar medium Results . For women with heavy group B Streptococcal colonization (more than 106 colony forming units/mL), the sensitivities were 36% (Quidel), 46% (ICON), and 100% (Strep B OIA). Specificities for all assays were high (98–100%), but variability was found in positive (79–100%) and negative (77–85%) predictive values. Although Strep B OIA reliably detects women with heavy group B Streptococcal colonization and is more sensitive than either the ICON or Quidel enzyme immunoassays, none of these rapid assays is sufficiently accurate for routine use in the intrapartum detection of women colonized with group B streptococcus.