How to make usage of the standardized EuroFlow 8-color protocols possible for instruments of different manufacturers
- PMID: 29154914
- DOI: 10.1016/j.jim.2017.11.007
How to make usage of the standardized EuroFlow 8-color protocols possible for instruments of different manufacturers
Abstract
A critical component of the EuroFlow standardization of leukemia/lymphoma immunophenotyping is instrument setup. Initially, the EuroFlow consortium developed a step-by-step standard operating protocol for instrument setup of ≥8-color flow cytometers that were available in 2006, when the EuroFlow activities started. Currently, there are 14 instruments from 9 manufacturers capable of 3-laser excitation and ≥8 color measurements. The specific adaptations required in the instrument set-up to enable them to acquire the standardized 8-color EuroFlow protocols are described here. Overall, all 14 instruments can be fitted with similar violet, blue and red lasers for simultaneous measurements of ≥8 fluorescent dyes. Since individual instruments differ both on their dynamic range (scale) and emission filters, it is not accurate to simply recalculate the target values to different scale, but adjustment of PMT voltages to a given emission filter and fluorochrome, is essential. For this purpose, EuroFlow has developed an approach using Type IIB (spectrally matching) particles to set-up standardized and fully comparable fluorescence measurements, in instruments from different manufacturers, as demonstrated here for the FACSCanto II, and Navios and MACSQuant flow cytometers. Data acquired after such adjustment on any of the tested cytometry platforms could be fully superimposed and therefore analyzed together. The proposed approach can be used to derive target values for any combination of spectrally distinct fluorochromes and any distinct emission filter of any new flow cytometry platform, which enables the measurement of the 8-color EuroFlow panels in a standardized way, by creating superimposable datafiles.
Keywords: EuroFlow; Flow cytometry; Fluorescent spectrum; Fluorochromes; Standardization.
Copyright © 2017. Published by Elsevier B.V.
Similar articles
-
FACSCanto II and LSRFortessa flow cytometer instruments can be synchronized utilizing single-fluorochrome-conjugated surface-dyed beads for standardized immunophenotyping.J Clin Lab Anal. 2020 Sep;34(9):e23361. doi: 10.1002/jcla.23361. Epub 2020 May 20. J Clin Lab Anal. 2020. PMID: 32430992 Free PMC article.
-
Comments on EuroFlow standard operating procedures for instrument setup and compensation for BD FACS Canto II, Navios and BD FACS Lyric instruments.J Immunol Methods. 2019 Dec;475:112680. doi: 10.1016/j.jim.2019.112680. Epub 2019 Oct 23. J Immunol Methods. 2019. PMID: 31655051
-
Reproducibility of Flow Cytometry Through Standardization: Opportunities and Challenges.Cytometry A. 2020 Feb;97(2):137-147. doi: 10.1002/cyto.a.23901. Epub 2019 Oct 8. Cytometry A. 2020. PMID: 31593368 Review.
-
Standardization of 8-color flow cytometry across different flow cytometer instruments: A feasibility study in clinical laboratories in Switzerland.J Immunol Methods. 2019 Dec;475:112348. doi: 10.1016/j.jim.2017.07.013. Epub 2017 Jul 29. J Immunol Methods. 2019. PMID: 28760670
-
EuroFlow standardization of flow cytometer instrument settings and immunophenotyping protocols.Leukemia. 2012 Sep;26(9):1986-2010. doi: 10.1038/leu.2012.122. Leukemia. 2012. PMID: 22948490 Free PMC article. Review.
Cited by
-
Commercial DURAClone panels for extending the repertoire of multicolour immunophenotypic panels in an academic flow cytometry laboratory in South Africa.Afr J Lab Med. 2022 Nov 29;11(1):1720. doi: 10.4102/ajlm.v11i1.1720. eCollection 2022. Afr J Lab Med. 2022. PMID: 36483322 Free PMC article.
-
CD20 Expression as a Possible Novel Prognostic Marker in CLL: Application of EuroFlow Standardization Technique and Normalization Procedures in Flow Cytometric Expression Analysis.Cancers (Basel). 2022 Oct 7;14(19):4917. doi: 10.3390/cancers14194917. Cancers (Basel). 2022. PMID: 36230840 Free PMC article.
-
The Euroflow PID Orientation Tube in the diagnostic workup of primary immunodeficiency: Daily practice performance in a tertiary university hospital.Front Immunol. 2022 Sep 13;13:937738. doi: 10.3389/fimmu.2022.937738. eCollection 2022. Front Immunol. 2022. PMID: 36177024 Free PMC article.
-
Evaluation of fixed-panel, multicolour ClearLLab 10C at an academic flow cytometry laboratory in Johannesburg, South Africa.Afr J Lab Med. 2022 Jul 15;11(1):1458. doi: 10.4102/ajlm.v11i1.1458. eCollection 2022. Afr J Lab Med. 2022. PMID: 35937760 Free PMC article.
-
Impact of Pre-Analytical and Analytical Variables Associated with Sample Preparation on Flow Cytometric Stainings Obtained with EuroFlow Panels.Cancers (Basel). 2022 Jan 18;14(3):473. doi: 10.3390/cancers14030473. Cancers (Basel). 2022. PMID: 35158741 Free PMC article.
Publication types
MeSH terms
LinkOut - more resources
Full Text Sources
Other Literature Sources