Orexin-A measurement in narcolepsy: A stability study and a comparison of LC-MS/MS and immunoassays

Orexin-A and -B are neuropeptides involved in sleep–wake regulation. In human narcolepsy type 1, this cycle is disrupted due to loss of orexin-producing neurons the hypothalamus. Cerebrospinal fluid (CSF) orexin-A measurement used diagnosis 1. Currently available immunoassays may lack specificity for accurate orexin quantification. We developed validated a liquid chromatography mass spectrometry assay (LC-MS/MS) CSF B. samples from 1 (n = 22) 2 6) non-narcoleptic controls 44). Stable isotope-labeled internal standards were added before solid-phase extraction quantification by LC-MS/MS. The also assayed commercial radioimmunoassay (RIA, n 42) enzymatic immunoassay (EIA, 72) kits. Stability orexins was studied 12 months. Our has good sensitivity (10 pmol/L 35 pg/mL) wide linear range (35–3500 pg/mL). Added stable 3 months, respectively, when frozen. median concentration patients <35 pg/mL (range < 35–131 pg/mL), which lower than that control individuals (98 pg/mL, 35–424 concentrations determined using our LC-MS/MS five times those measured with RIA. Orexin-B undetectable. as compared controls. RIA yielded on average higher