Resolution of RHCE Haplotype Ambiguities in Transfusion Settings

Red blood cell (RBC) transfusion, limited by patient alloimmunization, demands accurate blood group typing. The Rh system requires specific attention due to the limitations of serological phenotyping methods. Although these have been compensated for by molecular biology solutions, some RhCE ambiguit...

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Bibliographic Details
Published in:International journal of molecular sciences 2024-05, Vol.25 (11), p.5868
Main Authors: Izard, Caroline, Laget, Laurine, Beley, Sophie, Bichel, Nelly, De Boisgrollier, Lugdivine, Picard, Christophe, Chiaroni, Jacques, Di Cristofaro, Julie
Format: Article
Language:English
Subjects:
Alleles
Ambiguity
Antigens
Blood groups
Blood Transfusion - methods
Ethylenediaminetetraacetic acid
Genotype & phenotype
Haplotypes
Hematopoietic stem cells
Humans
Life Sciences
Molecular biology
Phenotype
Polymorphism
Reticulocytes - metabolism
Rh-Hr Blood-Group System - genetics
RNA, Messenger - genetics
Serology
Sickle cell anemia
Transplantation
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Summary:Red blood cell (RBC) transfusion, limited by patient alloimmunization, demands accurate blood group typing. The Rh system requires specific attention due to the limitations of serological phenotyping methods. Although these have been compensated for by molecular biology solutions, some RhCE ambiguities remain unresolved. The mRNA length is compatible with full-length analysis and haplotype discrimination, but the mRNA analyses reported so far are based on reticulocyte isolation and molecular biology protocols that are fastidious to implement in a routine context. We aim to present the most efficient reticulocyte isolation method, combined with an RT-PCR sequencing protocol that embraces the phasing of all haplotype configurations and identification of any allele. Two protocols were tested for reticulocyte isolation based either on their size/density properties or on their specific antigenicity. We show that the reticulocyte sorting method by antigen specificity from EDTA blood samples collected up to 48 h before processing is the most efficient and that the combination of an -specific RT-PCR followed by allele-specific sequencing enables analysis of cDNA haplotypes. All samples analyzed show full concordance between phenotype and haplotype sequencing. Two samples from the immunohematology laboratory with ambiguous results were successfully analyzed and resolved, one of them displaying a novel allele ( *03 c.340C>T).
ISSN:1422-0067
1661-6596
1422-0067
DOI:10.3390/ijms25115868