Systematic RH genotyping and variant identification in French donors of African origin.

BACKGROUND: RH molecular analysis has enabled the documentation of numerous variants of RHD and RHCE alleles, especially in individuals of African origin. The aim of the present study was to determine the type and frequency of D and/or RhCE variants among blood donors of African origin in France, by performing a systematic RH molecular analysis, in order to evaluate the implications for blood transfusion of patients of African origin. MATERIALS AND METHODS: Samples from 316 African blood donors, whose origin was established by their Fy(a-b-) phenotype, were first analysed using the RHD and RHCE BeadChips Kit (BioArray Solutions, Immucor, Warren, NJ, USA). Sequencing was performed when necessary. RESULTS: RHD molecular analysis showed that 26.2% of donors had a variant RHD allele. It allowed the prediction of a partial D in 11% of cases. RHCE molecular analysis showed that 14.2% of donors had a variant RHCE allele or RH [RN or (C)ces] haplotype. A rare Rh phenotype associated with the loss of a high-prevalence antigen or partial RhCE antigens were predicted from RHCE molecular analysis in 1 (0.3%) and 17 (5%) cases, respectively. DISCUSSION: Systematic RHD and RHCE molecular analysis performed in blood donors of African origin provides transfusion-relevant information for individuals of African origin because of the frequency of variant RH alleles. RH molecular analysis may improve transfusion therapy of patients by allowing better donor and recipient matching, based not only on phenotypically matched red blood cell units, but also on units that are genetically matched with regards to RhCE variants.

Anti-D investigations in individuals expressing weak D Type 1 or weak D Type 2: allo- or autoantibodies?

BACKGROUND: Whether anti-D produced by individuals with a weak D phenotype are allo- or autoantibodies remains a matter of debate even though blood transfusion practice is impacted. The aim of our study was to determine the serologic features of anti-D in individuals expressing the most frequent weak D type in Caucasians that are weak D Type 1 or weak D Type 2, to assess whether anti-D were allo- or autoantibodies. STUDY DESIGN AND METHODS: Serologic D typing and molecular analysis enabled the including of 121 weak D Type 1 individuals and 99 weak D Type 2 individuals in our study. Serologic features of anti-D included autologous controls, direct antiglobulin test, elution, and titration of anti-D before and after adsorption of serum on autologous red blood cells (RBCs). RESULTS: Serologic D typing showed a variable reactivity of RBCs expressing weak D Type 1 or weak D Type 2 (4+ to 0). Anti-D was identified in six weak D Type 1 and six weak D Type 2 individuals, respectively. The serologic data were in favor of autoantibodies. CONCLUSION: A complete anti-D investigation in individuals with a D variant (weak D or partial D identified by molecular analysis) should be systematically performed before any valid conclusion on the nature of the antibody. Transfusing weak D Type 1 or weak D Type 2 patients with D+ RBC units should be recommended. Weak D Type 1 or weak D Type 2 pregnant women do not need anti-D immunoprophylaxis.

Analysis of RhCE variants among 806 individuals in France: considerations for transfusion safety, with emphasis on patients with sickle cell disease.

BACKGROUND: DNA testing has enabled the documenting of numerous variants of RHCE alleles, especially in individuals of African origin. The risk for production of clinically significant alloantibodies to Rh antigens of patients carrying variant RHCE alleles has led us to analyze the different RhCE variants investigated by molecular biology. Alloimmunization was analyzed regarding the RHCE genetic profile. STUDY DESIGN AND METHODS: Samples from 806 individuals with altered expression of RhCE antigens and/or producing anti-RhCE in the presence of the corresponding antigen were analyzed. RESULTS: A total of 572 individuals were shown to express RhCE variants. Variant RHCE*ce alleles and RH haplotypes were identified in 83% of cases, the most frequent ones being the R(N) haplotype, the ceMO allele, the (C)ce(s) haplotype/ce(s) 1006 allele, and the ceAR allele identified in 36, 23, 20, and 17% of the tested samples, respectively. The absence of a high-prevalence Rh antigen was documented in 93 individuals. Partial C and partial e were expressed by 53% of individuals with RhCE variants. Rh antibodies were identified in 127 (20%) of 623 patients. They were found to be alloantibodies in 48 (38%) of these 127 patients. Alloimmunization against a high-prevalence Rh antigen was detected in 25% of cases. CONCLUSION: The challenge in clinical red blood cell (RBC) transfusion of patients with sickle cell disease, notably, would be to provide not only phenotypically matched, but also genetically matched, RBC units regarding RhCE variants.

Blood group genotyping by high-throughput DNA analysis applied to 356 reagent red blood cell samples.

BACKGROUND: DNA analysis for the prediction of blood group antigen expression has broad implications in transfusion medicine. It may be of particular interest especially to detect variants, when antigen expression is weak or altered. The use of high-throughput DNA analysis has never been applied to donors whose red blood cells (RBCs) are selected for reagent RBCs. The aim of this study was to analyze the concordance between the serologic phenotype and that predicted from DNA analysis in panel donors, to determine the benefit of the use of DNA analysis in reagent RBC selection strategy. STUDY DESIGN AND METHODS: The "Panel National de Référence du Centre National de Référence sur les Groupes Sanguins" is a reference reagent mainly used for antibody identification. DNA genotyping of 356 panel donors was performed with BeadChips (human erythrocyte antigen v1.2 BeadChips, BioArray Solutions). The comparison between serologic phenotype and that predicted from DNA analysis held on 8876 paired results obtained from 10 blood group systems and 25 antigens. RESULTS: A 99.95% concordance was observed. Discrepancies in four cases (RH, KEL, LU, and DO systems) were analyzed. Genotyping precisions on the Duffy system were of particular interest. No new rare blood group was observed. CONCLUSION: Systematic DNA analysis of panel donors should unquestionably change the management of reagent RBC selection. The notion of "antigens in double dose" should evolve regarding data obtained from DNA analysis, allowing an improved quality of reagent RBCs for antibody screening and identification.

Anti-HrB and anti-hrb revisited.

BACKGROUND: Since their description in the 1970s, anti-Hr(B) (antibody against a high-prevalence Rh antigen) and anti-hr(B) (anti-e-like antibody) are still a subject of debate about representing two aspects of a global immune response or being two independent antibodies. STUDY DESIGN AND METHODS: The aim of this study was to evaluate the immune response against the antigens of Rh system of 30 individuals presenting a hr(B)(RH31)- phenotype. Genomic analysis of RH genes was performed in all individuals. RESULTS: Among the 30 individuals, 27 had a Hr(B)(RH34)- phenotype. No immunization against Rh antigens was found in 16 individuals. Three individuals made anti-D only, whereas six individuals made anti-Hr(B) (four with anti-hr(B) and two without anti-hr(B)) and two individuals made anti-hr(B) without anti-Hr(B). Among the 30 individuals, three had a Hr(B)+ phenotype. No immunization against Rh antigens was found in one individual, whereas two individuals made anti-hr(B); the genomic analysis of selected individuals showed the presence of a (C)ce(s) haplotype, either Type 1 or Type 2, and a DIII Type 5 ce(s) haplotype, in the homozygous state, in compound heterozygosity with each other or in heterozygosity with a DcE haplotype. Genomic data were in accordance with serologic data. CONCLUSION: Our data provide the evidence that anti-Hr(B) and anti-hr(B) are independent antibodies, defining two different specificities. These antibodies may be produced by individuals expressing variants of RhCE protein. Serologic and molecular data indicate that e antigen encoded by the (C)ce(s) haplotype is a partial antigen. In individuals carrying a (C)ce(s) haplotype, the risk and the type of alloimmunization to Rh antigens are related to the second Rh haplotype.