title: Quality assessment program for EuroFlow protocols: summary results of four-year (2010-2013) quality assurance rounds.
abstract: Flow cytometric immunophenotyping has become essential for accurate diagnosis, classification, and disease monitoring in hemato-oncology. The EuroFlow Consortium has established a fully standardized "all-in-one" pipeline consisting of standardized instrument settings, reagent panels, and sample preparation protocols and software for data analysis and disease classification. For its reproducible implementation, parallel development of a quality assurance (QA) program was required. Here, we report on the results of four consecutive annual rounds of the novel external QA EuroFlow program. The novel QA scheme aimed at monitoring the whole flow cytometric analysis process (cytometer setting, sample preparation, acquisition and analysis) by reading the median fluorescence intensities (MedFI) of defined lymphocytes' subsets. Each QA participant applied the predefined reagents' panel on blood cells of local healthy donors. A uniform gating strategy was applied to define lymphocyte subsets and to read MedFI values per marker. The MedFI values were compared with reference data and deviations from reference values were quantified using performance score metrics. In four annual QA rounds, we analyzed 123 blood samples from local healthy donors on 14 different instruments in 11 laboratories from nine European countries. The immunophenotype of defined cellular subsets appeared sufficiently standardized to permit unified (software) data analysis. The coefficient of variation of MedFI for 7 of 11 markers performed repeatedly below 30%, average MedFI in each QA round ranged from 86 to 125% from overall median. Calculation of performance scores was instrumental to pinpoint standardization failures and their causes. Overall, the new EuroFlow QA system for the first time allowed to quantify the technical variation that is introduced in the measurement of fluorescence intensities in a multicentric setting over an extended period of time. EuroFlow QA is a proficiency test specific for laboratories that use standardized EuroFlow protocols. It may be used to complement, but not replace, established proficiency tests. © 2014 International Society for Advancement of Cytometry.
authors.affiliation: CLIP - Childhood Leukemia Investigation Prague and Department of Pediatric Hematology and Oncology, 2nd Faculty of Medicine, Charles University Prague and University Hospital Motol, Prague, Czech Republic.
authors.forename: Tomas
authors.initials: T
authors.lastname: Kalina
authors.forename: Juan
authors.initials: J
authors.lastname: Flores-Montero
authors.forename: Quentin
authors.initials: Q
authors.lastname: Lecrevisse
authors.forename: Carlos E
authors.initials: CE
authors.lastname: Pedreira
authors.forename: Vincent H J
authors.initials: VH
authors.lastname: van der Velden
authors.forename: Michaela
authors.initials: M
authors.lastname: Novakova
authors.forename: Ester
authors.initials: E
authors.lastname: Mejstrikova
authors.forename: Ondrej
authors.initials: O
authors.lastname: Hrusak
authors.forename: Sebastian
authors.initials: S
authors.lastname: Böttcher
authors.forename: Dennis
authors.initials: D
authors.lastname: Karsch
authors.forename: Łukasz
authors.initials: Ł
authors.lastname: Sędek
authors.forename: Amelie
authors.initials: A
authors.lastname: Trinquand
authors.forename: Nancy
authors.initials: N
authors.lastname: Boeckx
authors.forename: Joana
authors.initials: J
authors.lastname: Caetano
authors.forename: Vahid
authors.initials: V
authors.lastname: Asnafi
authors.forename: Paulo
authors.initials: P
authors.lastname: Lucio
authors.forename: Margarida
authors.initials: M
authors.lastname: Lima
authors.forename: Ana
authors.initials: A
authors.lastname: Helena Santos
authors.forename: Paola
authors.initials: P
authors.lastname: Bonaccorso
authors.forename: Alita J
authors.initials: AJ
authors.lastname: van der Sluijs-Gelling
authors.forename: Anton W
authors.initials: AW
authors.lastname: Langerak
authors.forename: Marta
authors.initials: M
authors.lastname: Martin-Ayuso
authors.forename: Tomasz
authors.initials: T
authors.lastname: Szczepański
authors.forename: Jacques J M
authors.initials: JJ
authors.lastname: van Dongen
authors.forename: Alberto
authors.initials: A
authors.lastname: Orfao
mesh_terms: D005060:Europe; D005434:Flow Cytometry / Q000379:methods; D064368:Healthy Volunteers; D016130:Immunophenotyping / Q000379:methods; D007938:Leukemia / Q000145:classification / Q000175:diagnosis; D016131:Lymphocyte Subsets / Q000276:immunology; D008223:Lymphoma / Q000145:classification / Q000175:diagnosis; D011786:Quality Control; D012015:Reference Standards; D012016:Reference Values; D006801:Humans
keywords: EuroFlow; flow cytometry; immunophenotyping; leukemia; lymphoma; quality assessment