B-lymphoblastoid cell lines as a source of reference DNA for human platelet and neutrophil antigen genotyping

Abstract

Background: Human platelet and neutrophil antigens (HPAs, HNAs) are targets for platelet or granulocyte antibodies causing immune thrombocytopenia or neutropenia, respectively. Currently, genotyping is replacing phenotyping as the preferred method of diagnosis of immune cytopenia. To establish a reliable genotyping analysis, however, the availability as reference DNA of genomic DNA from persons of known genotype is essential.

Study design and methods: By the use of Epstein-Barr virus transformation, panels of B-lympho-blastoid cell lines (B-LCLs) from HPA- and HNA-phenotyped individuals were developed. Genomic DNA was isolated from these cell lines and tested as reference DNA for genotyping of persons for HPAs and HNAs.

Results: DNA derived from these B-LCLs was typed by polymerase chain reaction-restriction fragment length polymorphism and -sequence-specific primers. The results were in accordance with the genotyping from peripheral blood cells. These results were confirmed by 24 laboratories in Germany in a blind study.

Conclusion: The inexhaustible source of reference DNA derived from B-LCLs allowed the evaluation of reliable HPA and HNA genotyping for quality control purposes. It should facilitate the development of DNA typing in blood centers and clinical laboratories.