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Background: Human leukocyte antigen (HLA) class I molecules are expressed on platelets and can represent a source of alloimmunization in recipients of platelet transfusions. HLA mismatch between donors and recipients may be associated with the induction of anti-HLA antibodies, which can culminate in refractoriness to platelet transfusions. In the present study we analyzed HLA allele group frequencies and HLA expression levels on human platelets from blood donors., Materials and Methods: Platelet-rich plasma was collected from 139 donors to monitor platelet HLA class I expression by flow cytometry. DNA from donors with high and low platelet HLA expression was used in the genotype studies. Frequencies of large and normal-sized platelet subpopulations were determined and HLA class I expression was studied. Mean platelet volume (MPV) and platelet large-cell ratio (P-LCR) were analyzed in both groups of donors., Results: The analysis showed variable platelet HLA class I expression with significant differences among donors. HLA class I allele group frequencies in donors with high and low platelet HLA expression showed distinctive genotypic features strictly related to expression level. The main allele groups found in samples with high platelet HLA class I expression were HLA-A*02, -A*68, -B*15, -B*49, and -C*03. Platelet HLA class I expression did not change over time or during freezing-thawing cycles. The analysis of platelet subpopulations showed a statistically significant higher expression of HLA class I molecules on large platelets than on normal-sized platelets. Moreover, donors with high HLA class I expression showed a higher frequency of large platelets (p<0.0001). The analysis of P-LCR in both groups of donors showed a statistically significant difference (p<0.05) within high HLA-expressing donors., Discussion: Our data suggest an allele-dependent expression of HLA class I molecules on human platelets with distinct HLA allele group frequencies and different platelet subpopulation frequencies among blood donors.