D variants in the population of D‐negative blood donors in the north‐eastern region of Croatia

Objectives The aim of this study was to determine RHESUS D GENE ( RHD) allelic variants among Croatian D‐negative blood donors and compare our results with respective data from other European countries. Background Altered or reduced D antigen expression can result in D variants, which can be mistyped and can lead to the alloimmunisation of the blood recipient. RHD genotyping can distinguish D variants: weak D, partial D and DEL, thus preventing alloimmunisation. Material/methods A total of 6523 samples obtained from D‐negative Croatian donors were screened for the presence of RHD using the real‐time polymerase chain reaction (PCR) method. PCR‐SSP was performed for D variant genotyping by using commercial genotyping kits (Inno‐Train, Kronberg, Germany). Genomic DNA sequencing for all 10 exons of the RHD was performed when the genotyping kits failed to assign a D variant. Results RHD molecular screening revealed 23 (0.35%) RHD ‐PCR positive samples, all C/E positive, in decreasing frequency: 11 hybrid RHD‐CE (2‐9) D‐CE variants, 4 weak partial D type 11 and 2 weak D type 2. Six samples remained unresolved and were sequenced. For 12 of 23 samples (excluding large hybrids), an adsorption/elution of anti‐D serum was performed, confirming that all 12 were RhD+. The calculated frequency of clinically significant D alleles in RhD‐negative blood donors was 1:543 (0.18%) or 1:53 (1.89%) in C/E blood donors. Conclusion Data on the significant frequency of D variants among serologically D‐negative blood donors in the north‐eastern region of Croatia could help in introducing RHD molecular screening of blood donors in a routine workflow.