Development of a Nonradioactive Platelet Serotonin Uptake and Release Assay by Micro-Liquid Chromatography Tandem Mass Spectrometry Using Minimal Blood Volume

Siaw Li Chan, Xin Yi, Emily Wysocki, Rachael Bridgman, Jocelyn Gutierrez, Krzysztof Mikrut, Edward Ki Yun Leung, Kiang Teck J. Yeo, Jonathan L. Miller

Research output: Contribution to journalArticle

Abstract

Objectives: Analysis of platelet functional responses to stimuli is presently quite limited with respect to measurement of dense granule secretion. We sought to develop a nonradioactive assay of stimulated serotonin release using liquid chromatography tandem mass spectrometry (LC-MS/MS). Methods: Citrated whole blood (200 μL) was incubated with deuterated serotonin (d45-HT). Following uptake by platelets, blood was diluted 10-fold and aliquots were incubated with platelet stimuli. Following stimulation, blood was further diluted, centrifuged, and supernatant was assayed for released d45-HT by micro-LC-MS/MS. Results: This study demonstrated a broad linear range of 50 to 2,000 pg/mL d45-HT, with a total precision of less than 15.0% coefficient of variation at all quality control levels and a limit of quantitation of 50 pg/mL. Conclusions: Quantification of d45-HT by micro-LC-MS/MS assay offers a highly sensitive, nonradioactive methodology for quantitating platelet serotonin uptake and dense granule secretion, requiring only small volumes of patient blood.

Original languageEnglish (US)
Pages (from-to)718-724
Number of pages7
JournalAmerican Journal of Clinical Pathology
Volume152
Issue number6
DOIs
StatePublished - Nov 4 2019

Keywords

  • Dense granule secretion
  • LC-MS/MS
  • Nonradioactive serotonin release assay
  • Platelet function

ASJC Scopus subject areas

  • Pathology and Forensic Medicine

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