High-resolution melting analysis as an alternative method for human neutrophil antigen genotyping

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Immunohematology

American National Red Cross

Subject: Medical Laboratory Technology

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ISSN: 0894-203X
eISSN: 1930-3955

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VOLUME 31 , ISSUE 1 (March 2015) > List of articles

High-resolution melting analysis as an alternative method for human neutrophil antigen genotyping

Kazuta Yasui / Mitsunobu Tanaka / Tomoya Hayashi / Nobuki Matsuyama / Ayumu Kuroishi / Rika. A. Furuta / Yoshihiko Tani / Fumiya Hirayama

Keywords : neutrophil, HNA, antibody, genotyping, highresolution melting analysis

Citation Information : Immunohematology. Volume 31, Issue 1, Pages 7-13, DOI: https://doi.org/10.21307/immunohematology-2019-064

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ABSTRACT

Human neutrophil antigen (HNA)-typed granulocyte panels are widely used to screen for the presence of HNA antibodies and to determine antibody specificity. Many laboratories screen donors for HNA genotypes using low-throughput methods such as allele-specific polymerase chain reaction (PCR), PCR–restriction fragment–length polymorphism, and multiplex PCR. In the present study, we used a high-resolution melting (HRM) analysis to determine HNA genotypes. For the HRM analysis, purified genomic DNA samples were amplified via PCR with HNA-specific primers. Nucleotide substitutions in genes encoding HNAs were differentiated on the basis of the HRM curves, and the results of HRM and DNA sequencing analyses were determined to be in complete agreement. The gene frequency of HNA-1a, -1b, -1c, -3a, -3b, -4a, -4b, -5a, and -5b in the Japanese population was consistent with the previous reports. Our results suggest that HRM analysis can be used for genotyping HNA antigens determined by single nucleotide substitutions.

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