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Full‐length recombinant choline transporter–like protein 2 containing arginine 154 reconstitutes the epitope recognized by HNA‐3a antibodies
Author(s) -
Kanack Adam J.,
Peterson Julie A.,
Sullivan Mia J.,
Bougie Daniel W.,
Curtis Brian R.,
Aster Richard H.
Publication year - 2012
Publication title -
transfusion
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.045
H-Index - 132
eISSN - 1537-2995
pISSN - 0041-1132
DOI - 10.1111/j.1537-2995.2011.03411.x
Subject(s) - epitope , antibody , hek 293 cells , context (archaeology) , transfection , microbiology and biotechnology , recombinant dna , chemistry , arginine , antigen , biology , biochemistry , immunology , gene , amino acid , paleontology
BACKGROUND: Recent reports have shown that the HNA‐3a leukocyte antigen, a target for antibodies that cause severe transfusion‐related acute lung injury, correlates with an arginine 154 (rather than glutamine) polymorphism in choline transporter–like protein 2 (CTL2) but did not show directly that R154 determines HNA‐3a. CTL2 peptides containing R154 are recognized by only half of HNA‐3a antibodies studied to date. Constructs that react with all HNA‐3a antibodies are needed to fully define the HNA‐3a epitope. STUDY DESIGN AND METHODS: HEK293 cells were transfected with cDNA encoding full‐length CTL2 linked to green fluorescent protein (GFP). Transfectants were selected for GFP expression and tested with antibodies specific for HNA‐3a and ‐3b. RESULTS: Each of 20 HNA‐3a antibodies reacted preferentially with HEK293 cells expressing the R154 CTL2 construct. An HNA‐3b antibody reacted only with CTL2 (Q154). CONCLUSIONS: These findings provide direct evidence that R154 in the context of full‐length CTL2 is both necessary and sufficient to create the HNA‐3a epitope but suggest that posttranslational modifications of the protein, for example, S‐S bonds or addition of glycans, are necessary for recognition of HNA‐3a by many antibodies. This could complicate development of an assay for large‐scale screening of blood donors to detect anti‐HNA‐3a.