Serological and genetic analysis of a B3 phenotype caused by c.259G > T in the ABO gene.

Abstract

Background: Mixed agglutination is a serological pattern in some ambiguous ABO blood type identification. This study focused on the serological and molecular genetic characteristics of a B3 phenotype induced by a c.259G > T mutation in the ABO gene.

Study design and methods: Serological methods such as gel cards and tubes were used to identify the ABO blood type of the patient, with fluorescent PCR for ABO genotyping and Sanger sequencing for analysing the ABO exons. Protein 3D Structure was simulated and further analysed using SWISS-MODLE and PyMOL. Both the wild-type (VAL-87, ABO*B.01) and the mutant (p.Val87Leu) plasmids were transfected into Hela cells to assess the agglutination intensity of the transfected cells with anti-B antibodies.

Results: Serological testing showed weak expression of the B antigen and mixed agglutination with anti-B antibodies. ABO genotyping indicated the presence of a B allele, but exon sequencing revealed an additional c.259G > T mutation in exon 6 based on the ABO*B.01 allele. The simulated three-dimensional structures of the proteins showed increased steric hindrance with mutations, leading to a relatively loose structure. The transfected Hela cells with the mutant (p.Val87Leu) plasmid exhibited a significantly reduced agglutination intensity with anti-B antibodies.

Conclusion: Based on comprehensive serological, genetic, and simulation analyses, it is concluded that the c.259G > T mutation in exon 6 of the ABO*B.01 allele results in an amino acid change within the enzymatic active site. This alteration likely impacts protein stability and reduces B antigen expression, leading to the B3 subtype phenotype.