RHD Allelic Identification Among D−Brazilian Blood Donors as a Routine Test Using Pools of DNA

Abstract

RHD alleles leading to a reduced expression of D antigen of the red blood cell (RBC) surface may be erroneously typed as D- by serology and may cause anti-D immunizations when transfused to recipients. To determine the occurrence of such alleles among apparent D- blood donors, molecular typing was implemented as a routine test using a pool of DNA. A total of 2,450 pretyped D- samples were tested in pools of 10 for the RHD-specific polymorphism in intron 4 and exon 7. Samples in polymer chain reaction (PCR) positive pools were individually reevaluated by exon-specific PCRs, sequencing, and serologic methods. Among 2,450 serologically D- blood donor samples tested, 101 (4.1%) carried the RHD gene. Nonfunctional RHD (RHDψ, RHD*CE(2-9)-D, and RHD*CE(3-7)-D), different weak D alleles such as RHD*weak D type 1, RHD*weak D type 4.3, RHD*weak D type 5, RHD*weak D type 38, and RHD*DEL were identified. We employed a PCR-based assay for RHD as a routine test using pools of ten DNA blood donor samples. The integration of RHD genotyping into the routine screening program using pools of DNA samples was straightforward. As a consequence, 19 (0.8%) blood donors carrying a weak D and Del phenotypes with the potential of causing anti-D immunizations in recipients were reclassified as D+. For each population, it would be necessary to adapt the RHD genotyping strategy to the spectrum of prevalent alleles.