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Comparative analysis of serotonin in equine plasma with liquid chromatography–tandem mass spectrometry and enzyme-linked immunosorbent assay

Sara Torfs (UGent) , An Maes (UGent) , Catherine Delesalle (UGent) , Piet Deprez (UGent) and Siska Croubels (UGent)
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Abstract
Serotonin is regularly measured in equine platelet-poor plasma in research settings. However, reported reference values vary between studies, partially because plasma serotonin concentrations are very low and a reliable and affordable detection method is lacking. A simple, rapid, and sensitive method for serotonin determination in equine platelet-poor plasma using liquid chromatography–tandem mass spectrometry (LC-MS/MS) was developed and validated. Results of a commercially available enzyme-linked immunosorbent assay (ELISA) were compared to the LC-MS/MS results, in order to validate a test more suitable for use in a clinical situation. For LC-MS/MS, 500 µl of plasma was required, and deuterated serotonin was used as an internal standard. The sample preparation was based upon a simple liquid extraction into ethyl acetate. Chromatographic separation was performed with an acetic acid–acetonitrile mobile phase gradient elution. Linearity was demonstrated between 3 ng/ml and 100 ng/ml. A limit of quantification of 3 ng/ml was achieved, corresponding to a limit of detection of 0.10 ng/ml. Comparison of LC-MS/MS and ELISA with Passing–Bablok regression and Bland–Altman plotting showed a poor agreement between the 2 methods, with an increasing difference within the higher range of measurements. Caution is needed when extrapolating results from sources using different analytical techniques.
Keywords
horses, serotonin, liquid chromatography, Enzyme-linked immunosorbent assay, tandem mass spectrometry

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Chicago
Torfs, Sara, An Maes, Catherine Delesalle, Piet Deprez, and Siska Croubels. 2012. “Comparative Analysis of Serotonin in Equine Plasma with Liquid Chromatography–tandem Mass Spectrometry and Enzyme-linked Immunosorbent Assay.” Journal of Veterinary Diagnostic Investigation 24 (6): 1035–1042.
APA
Torfs, S., Maes, A., Delesalle, C., Deprez, P., & Croubels, S. (2012). Comparative analysis of serotonin in equine plasma with liquid chromatography–tandem mass spectrometry and enzyme-linked immunosorbent assay. JOURNAL OF VETERINARY DIAGNOSTIC INVESTIGATION, 24(6), 1035–1042.
Vancouver
1.
Torfs S, Maes A, Delesalle C, Deprez P, Croubels S. Comparative analysis of serotonin in equine plasma with liquid chromatography–tandem mass spectrometry and enzyme-linked immunosorbent assay. JOURNAL OF VETERINARY DIAGNOSTIC INVESTIGATION. 2012;24(6):1035–42.
MLA
Torfs, Sara, An Maes, Catherine Delesalle, et al. “Comparative Analysis of Serotonin in Equine Plasma with Liquid Chromatography–tandem Mass Spectrometry and Enzyme-linked Immunosorbent Assay.” JOURNAL OF VETERINARY DIAGNOSTIC INVESTIGATION 24.6 (2012): 1035–1042. Print.
@article{3062408,
  abstract     = {Serotonin is regularly measured in equine platelet-poor plasma in research settings. However, reported reference values vary between studies, partially because plasma serotonin concentrations are very low and a reliable and affordable detection method is lacking. A simple, rapid, and sensitive method for serotonin determination in equine platelet-poor plasma using liquid chromatography--tandem mass spectrometry (LC-MS/MS) was developed and validated. Results of a commercially available enzyme-linked immunosorbent assay (ELISA) were compared to the LC-MS/MS results, in order to validate a test more suitable for use in a clinical situation. For LC-MS/MS, 500 {\textmu}l of plasma was required, and deuterated serotonin was used as an internal standard. The sample preparation was based upon a simple liquid extraction into ethyl acetate. Chromatographic separation was performed with an acetic acid--acetonitrile mobile phase gradient elution. Linearity was demonstrated between 3 ng/ml and 100 ng/ml. A limit of quantification of 3 ng/ml was achieved, corresponding to a limit of detection of 0.10 ng/ml. Comparison of LC-MS/MS and ELISA with Passing--Bablok regression and Bland--Altman plotting showed a poor agreement between the 2 methods, with an increasing difference within the higher range of measurements. Caution is needed when extrapolating results from sources using different analytical techniques.},
  author       = {Torfs, Sara and Maes, An and Delesalle, Catherine and Deprez, Piet and Croubels, Siska},
  issn         = {1040-6387},
  journal      = {JOURNAL OF VETERINARY DIAGNOSTIC INVESTIGATION},
  language     = {eng},
  number       = {6},
  pages        = {1035--1042},
  title        = {Comparative analysis of serotonin in equine plasma with liquid chromatography--tandem mass spectrometry and enzyme-linked immunosorbent assay},
  url          = {http://dx.doi.org/10.1177/1040638712457928},
  volume       = {24},
  year         = {2012},
}

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