A1,2BO1,2 genotyping by multiplexed allele‐specific PCR

The ABO blood group is the most clinically important human alloantigen system in transfusion medicine. The system involves three antigens A, B and H. H antigen is converted to either A or B by the activity of alpha1-->3-N-acetyl-galactosaminyl transferase (A transferase) or alpha1-->3 galactosyl transferase (B transferase). The O phenotype is the result of an inactive glycosyltransferase, which is unable to glycosylate the H antigen. The immunological properties of the ABO system were identified at the turn of the century; however, the genetic basis of the ABO system has only recently been characterized. This has enabled the development of a number of molecular ABO typing methods. Described here is a two-reaction multiplex allele-specific PCR (ASPCR) genotyping assay for the A1, A2, B, O1 and O2 subtypes. 11 different allele-specific oligonucleotide primers were selected to detect the presence or absence of the O1 associated G--> (-) deletion at base 261, the O2 associated G --> A substitution at base 802, the B associated G --> A substitution at base 803, and finally the A2 associated C --> (-) deletion at base 1059. A total of 122 peripheral blood samples were genotyped and serologically forward and reverse typed. A concordance rate of 98.4% (120/122 samples) was observed between the actual genotype and the serologically-based predicted genotype. These results indicate that this assay provides a rapid, accurate, and simple method for A(1,2)BO(1,2) genotyping that serves as a useful supplement to standard serological ABO typing.