RAPID RH-D GENOTYPING BY POLYMERASE CHAIN REACTION-BASED AMPLIFICATION OF DNA

Rh (rhesus) D is the dominant antigen of the ph blood group system. Re cent advances in characterization of the nucleotide sequence of the cD NA(s) encoding the Rh D polypeptide allow the determination of the Rh D genotype at the DNA level. This can be of help in cases in which red blood cells are not available for phenotyping, eg, when it concerns a fetus. We have tested three independent DNA typing methods based on t he polymerase chain reaction [PCR) for their suitability to determine the Rh D genotype. DNA derived from peripheral blood mononuclear cells from 234 Rh-phenotyped healthy donors (178 Rh D positive and 56 Rh D negative) was used in the PCR. The Rh D genotypes, as determined with a method based on the allele-specific amplification of the 3' noncodin g region of the Rh D gene described by Bennett et al (N Engl J Med 329 :607, 1993), were not concordant with the serologically established ph enotypes in all cases. We have encountered 5 discrepant results, ie, 3 false-positive and 2 false-negative (a father and child). ph D genoty ping with the second method was performed by PCR amplification of exon 7 of the D gene with allele-specific primers. In all donors phenotype d as D positive tested so far (n = 178), the results of molecular geno typing with this method were concordant with the serologic results, wh ereas a false-positive result was obtained in one of the D-negative do nors (also false-positive in the first method). Complete agreement was found between genotypes determined in the third method, based on a 60 0-bp deletion in intron 4 of the Rh D gene described by Arce et al (Bl ood 82:651, 1993), and serologically determined phenotypes. The Rh blo od group system is complex, and unknown polymorphisms at the DNA level are expected to exist. Therefore, although genotypes determined by th e method of Arce et al were in agreement with serotypes, it cannot yet be regarded as the golden standard. More experience with this or othe r methods is still needed. (C) 1995 by The American Society of Hematol ogy.