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A high‐performance liquid chromatography method for the serotonin release assay is equivalent to the radioactive method
Author(s) -
SonoKoree N. K.,
Crist R. A.,
Frank E. L.,
Rodgers G. M.,
Smock K. J.
Publication year - 2016
Publication title -
international journal of laboratory hematology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.705
H-Index - 55
eISSN - 1751-553X
pISSN - 1751-5521
DOI - 10.1111/ijlh.12442
Subject(s) - high performance liquid chromatography , chromatography , serotonin , chemistry , platelet , reagent , biochemistry , medicine , receptor
Summary Introduction The serotonin release assay ( SRA ) is considered the gold standard laboratory test for heparin‐induced thrombocytopenia ( HIT ). The historic SRA method uses platelets loaded with radiolabeled serotonin to evaluate platelet activation by HIT immune complexes. However, a nonradioactive method is desirable. We report the performance characteristics of a high‐performance liquid chromatography ( HPLC ) SRA method. Methods We validated the performance characteristics of an HPLC ‐ SRA method, including correlation with a reference laboratory using the radioactive method. Serotonin released from reagent platelets was quantified by HPLC using fluorescent detection. Results were expressed as % release and classified as positive, negative, or indeterminate based on previously published cutoffs. Results Serum samples from 250 subjects with suspected HIT were tested in the HPLC ‐ SRA and with the radioactive method. Concordant classifications were observed in 230 samples (92%). Sera from 41 healthy individuals tested negative. Between‐run imprecision studies showed standard deviation of <6 (% release) for positive, weak positive, and negative serum pools. Stability studies demonstrated stability after two freeze–thaw cycles or up to a week of refrigeration. Conclusion The HPLC ‐ SRA has robust performance characteristics, equivalent to the historic radioactive method, but avoids the complexities of working with radioactivity.