Abstract The molecular background of variant forms of GYPB is not well studied in Brazilians of African descent. The present study was carried out to determine the molecular bases of the S–s– phenotype and the frequency of GYPB*S silent gene for the S–s+ phenotype in a blood donor population of African Brazilians. In this study, 165 blood samples from African Brazilians (Northeastern Brazil) who phenotyped as S–s– (n = 17) and S–s+ (n = 148) by hemagglutination were selected. Allele-specific (AS)-PCR and PCR-restriction fragment length polymorphism (RFLP) were used to identify the variant forms of GYPB. In 13 of 17 S–s– samples (76.5%), both GYPB were deleted. In 137 of the 148 S–s+ samples (92.6%), the AS-PCR was consistent with the S–s+ phenotype. In 4 of the S–s– samples (23.5%) and 11 of the S–s+ samples (7.4%), the AS-PCR showed the presence of a GYPB*S allele associated with silencing of S. In the 4 donors with the S–s– phenotype, there was homozygosity (or hemizygosity) for the GYP(P2) allele (n = 2), homozygosity (or hemizygosity) for the GYP(NY) allele (n = 1), and heterozygosity for the GYP(P2) and GYP(NY) alleles (n = 1). In the 11 donors with the S–s+ phenotype, there was heterozygosity for GYP(P2) allele (n = 8) and heterozygosity for GYP(NY) allele (n = 3). This study reports for the first time the molecular mechanisms responsible for the S–s– phenotype in a population of African Brazilians and provides new information about the frequency and molecular bases of the GYPB*S silent gene (7.4%) in this population. Immunohematology 2008;24:148-153.