The platelet-specific alloantigen PlA1 (HPA-1a): a comparison of flow cytometric immunophenotyping and genotyping using polymerase chain reaction and restriction fragment length polymorphism in a Swedish blood donor population.
BACKGROUND
There is an increasing interest in the development of rapid and reliable techniques for platelet alloantigen typing.


METHODS
By use of standardized flow cytometry and a specific human alloantiserum, 236 Swedish blood donors were immunophenotyped for the platelet-specific alloantigen, PlA1 (HPA-1a).


RESULTS
Ten individuals (4.2%) had low fluorescence intensities and were considered PlA1-negative (HPA-1a-negative); all of them also demonstrated a PlA2/PlA2 (HPA-1b/1b) genotype in a polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP) assay of the underlying DNA polymorphism. The remaining population had clear positive fluorescence and was regarded as PlA1-positive (HPA-1a-positive). The fluorescence distribution histogram among PlA1-positive (HPA-1a-positive) individuals was dome-shaped, and those individuals who were homozygous for PlA1 (HPA-1a) could not be distinguished from those who were heterozygous. This finding was further substantiated by PCR-RFLP analysis of the PlA1/PlA2 (HPA-1a/1b) genotype; a heterozygous genotype was found among those having a medium fluorescence intensity as well as among those having a strong fluorescence intensity.


CONCLUSIONS
Flow cytometry is a valuable tool for large-scale detection of PlA1 (HPA-1a). However, flow cytometry based on only one antiserum cannot distinguish between homozygous and heterozygous carriers of PlA1 (HPA-1a). For zygosity testing and when platelets are difficult to obtain, the PCR-RFLP technique is the assay of choice.
Forsberg
B
B
Institution for Biomedical Laboratory Science, Sahlgrenska Hospital, University of Göteborg, Sweden.
Jacobsson
S
S
Stockelberg
D
D
Kutti
J
J
Rydberg
L
L
Wadenvik
H
H
D016824:Antigens, Human Platelet / Q000235:genetics* / Q000276:immunology*; D001483:Base Sequence; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D005838:Genotype; D006801:Humans; D016130:Immunophenotyping; D039661:Integrin beta3; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D013548:Sweden
D016824:Antigens, Human Platelet; C061248:ITGB3 protein, human; D039661:Integrin beta3