Centrally Determined Standardization of Flow Cytometry Methods Reduces Interlaboratory Variation in a Prospective Multicenter Study
| Autor: | Severine Vermeire, Silvio Danese, Geert R. D'Haens, Melissa Filice, Vipul Jairath, Michelle I. Smith, Hannelie Korf, Mark S. Silverberg, Jenny Jeyarajah, Stefania Vetrano, Azar Azad, Brian G. Feagan, David M. Spencer, Azucena Salas, Liset Westera, Dermot P.B. McGovern, William A. Faubion, Gijs R. van den Brink, Larry Stitt, Barrett G. Levesque, Lisa M. Shackelton, Niels Vande Casteele, Julián Panés, Tanja van Viegen, William J. Sandborn, Jonathan Cremer, Lars Eckmann, Janine Bilsborough |
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| Přispěvatelé: | Westera, L, van Viegen, T, Jeyarajah, J, Azad, A, Bilsborough, J, van den Brink, Gr, Cremer, J, Danese, S, D'Haens, G, Eckmann, L, Faubion, W, Filice, M, Korf, H, Mcgovern, D, Panes, J, Salas, A, Sandborn, Wj, Silverberg, M, Smith, Mi, Vermeire, S, Vetrano, S, Shackelton, Lm, Stitt, L, Jairath, V, Levesque, Bg, Spencer, Dm, Feagan, Bg, Casteele, Nv, Gastroenterology and Hepatology, Tytgat Institute for Liver and Intestinal Research, Amsterdam Gastroenterology Endocrinology Metabolism |
| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
Oncology medicine.medical_specialty Pathology Science & Technology Gastroenterology & Hepatology medicine.diagnostic_test Standardization business.industry Original Contributions Gastroenterology HUMAN IMMUNOLOGY Flow cytometry 03 medical and health sciences 030104 developmental biology 0302 clinical medicine Multicenter study Internal medicine medicine ASSAYS business Life Sciences & Biomedicine 030215 immunology |
| Zdroj: | Clinical and translational gastroenterology, 8(11). Nature Publishing Group Clinical and Translational Gastroenterology |
| ISSN: | 2155-384X |
| Popis: | OBJECTIVES: Flow cytometry (FC) aids in characterization of cellular and molecular factors involved in pathologic immune responses. Although FC has potential to facilitate early drug development in inflammatory bowel disease, interlaboratory variability limits its use in multicenter trials. Standardization of methods may address this limitation. We compared variability in FC-aided quantitation of T-cell responses across international laboratories using three analytical strategies. METHODS: Peripheral blood mononuclear cells (PBMCs) were isolated from three healthy donors, stimulated with phorbol 12-myristate 13-acetate and ionomycin at a central laboratory, fixed, frozen, and shipped to seven international laboratories. Permeabilization and staining was performed in triplicate at each laboratory using a common protocol and centrally provided reagents. Gating was performed using local gating with a local strategy (LGLS), local gating with a central strategy (LGCS), and central gating (CG). Median cell percentages were calculated across triplicates and donors, and reported for each condition and strategy. The coefficient of variation (CV) was calculated across laboratories. Between-strategy comparisons were made using a two-way analysis of variance adjusting for donor. RESULTS: Mean interlaboratory CV ranged from 1.8 to 102.1% depending on cell population and gating strategy (LGLS, 4.4-102.1%; LGCS, 10.9-65.6%; CG, 1.8-20.9%). Mean interlaboratory CV differed significantly across strategies and was consistently lower with CG. CONCLUSIONS: Central gating was the only strategy with mean CVs consistently lower than 25%, which is a proposed standard for pharmacodynamic and exploratory biomarker assays |
| Databáze: | OpenAIRE |
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