title: A high-performance liquid chromatography method for the serotonin release assay is equivalent to the radioactive method.
abstract: BACKGROUND
  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.
  
  
  CONCLUSIONS
  The HPLC-SRA has robust performance characteristics, equivalent to the historic radioactive method, but avoids the complexities of working with radioactivity.
authors.affiliation: ARUP Laboratories Institute for Clinical and Experimental Pathology, Salt Lake City, UT, USA.
authors.forename: N K
authors.initials: NK
authors.lastname: Sono-Koree
authors.affiliation: ARUP Laboratories Institute for Clinical and Experimental Pathology, Salt Lake City, UT, USA.
authors.forename: R A
authors.initials: RA
authors.lastname: Crist
authors.affiliation: ARUP Laboratories Institute for Clinical and Experimental Pathology, Salt Lake City, UT, USA.|Department of Pathology, University of Utah Health Sciences Center, Salt Lake City, UT, USA.
authors.forename: E L
authors.initials: EL
authors.lastname: Frank
authors.affiliation: ARUP Laboratories Institute for Clinical and Experimental Pathology, Salt Lake City, UT, USA.|Department of Pathology, University of Utah Health Sciences Center, Salt Lake City, UT, USA.|Department of Internal Medicine, University of Utah Health Sciences Center, Salt Lake City, UT, USA.
authors.forename: G M
authors.initials: GM
authors.lastname: Rodgers
authors.affiliation: ARUP Laboratories Institute for Clinical and Experimental Pathology, Salt Lake City, UT, USA.|Department of Pathology, University of Utah Health Sciences Center, Salt Lake City, UT, USA.
authors.forename: K J
authors.initials: KJ
authors.lastname: Smock
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods* / Q000592:standards*; D006493:Heparin / Q000009:adverse effects*; D006801:Humans; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012701:Serotonin / Q000097:blood*; D013921:Thrombocytopenia / Q000139:chemically induced* / Q000175:diagnosis*
chemical_list: D012701:Serotonin; D006493:Heparin
keywords: Heparin-induced thrombocytopenia; high-performance liquid chromatography; laboratory testing; platelets; serotonin release assay