Standardizing Flow Cytometry Immunophenotyping Analysis from the Human ImmunoPhenotyping Consortium

被引：218

作者：

Finak, Greg ^{[1
]}

Langweiler, Marc ^{[2
]}

Jaimes, Maria ^{[3
]}

Malek, Mehrnoush ^{[4
]}

Taghiyar, Jafar ^{[4
]}

Korin, Yael ^{[5
]}

Raddassi, Khadir ^{[6
]}

Devine, Lesley ^{[6
]}

Obermoser, Gerlinde ^{[7
]}

Pekalski, Marcin L. ^{[8
]}

Pontikos, Nikolas ^{[8
]}

Diaz, Alain ^{[9
]}

Heck, Susanne ^{[10
]}

Villanova, Federica ^{[10
]}

Terrazzini, Nadia ^{[11
]}

Kern, Florian ^{[12
]}

Qian, Yu ^{[13
]}

Stanton, Rick ^{[13
]}

Wang, Kui ^{[14
]}

Brandes, Aaron ^{[15
]}

Ramey, John ^{[1
]}

Aghaeepour, Nima ^{[4
,16
]}

Mosmann, Tim ^{[17
]}

Scheuermann, Richard H. ^{[13
]}

Reed, Elaine ^{[5
]}

Palucka, Karolina ^{[7
]}

Pascual, Virginia ^{[7
]}

Blomberg, Bonnie B. ^{[9
]}

Nestle, Frank ^{[10
]}

Nussenblatt, Robert B. ^{[18
]}

Brinkman, Ryan Remy ^{[4
,19
]}

Gottardo, Raphael ^{[1
]}

Maecker, Holden ^{[20
]}

Mccoy, J. Philip ^{[21
]}

机构：

[1] Fred Hutchinson Canc Res Ctr, Vaccine & Infect Dis Div, Seattle, WA 98109 USA

[2] NIH, Hematol Branch, Bldg 10, Bethesda, MD 20892 USA

[3] BD Biosci, San Jose, CA USA

[4] British Columbia Canc Agcy, Terry Fox Lab, Vancouver, BC V3J 4W6, Canada

[5] UCLA Pathol & Lab Med, Los Angeles, CA USA

[6] Yale Univ, Sch Med, Dept Neurol, New Haven, CT USA

[7] Baylor Inst Immunol Res, Dallas, TX USA

[8] Univ Cambridge, Cambridge Inst Med Res, JDRF Wellcome Trust Diabet & Inflammat Lab, Cambridge, England

[9] Univ Miami, Miller Sch Med, Dept Microbiol & Immunol, Miami, FL 33136 USA

[10] Guys Hosp, Guys & St Thomas Hosp, London SE1 9RT, England

[11] Univ Brighton, Sch Pharm & Biomol Sci, Brighton BN2 4GJ, E Sussex, England

[12] Brighton & Sussex Med Sch, Div Med, Brighton BN1 9PS, E Sussex, England

[13] J Craig Venter Inst, Dept Informat, La Jolla, CA 92037 USA

[14] Univ Queensland, Sch Math & Phys, Brisbane, Qld, Australia

[15] Broad Inst MIT & Harvard, Cambridge, MA 02142 USA

[16] Stanford Univ, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA

[17] Univ Rochester, Med Ctr, Sch Med & Dent, Rochester, NY 14642 USA

[18] NEI, Immunol Lab, NIH, Bldg 10, Bethesda, MD 20892 USA

[19] Univ British Columbia, Dept Med Genet, Vancouver, BC V5Z 1M9, Canada

[20] Stanford Univ, Sch Med, Inst Immun Transplantat & Infect, Stanford, CA 94305 USA

[21] NHLBI Flow Cytometry Core, NIH, Bethesda, MD 20892 USA

来源：

SCIENTIFIC REPORTS | 2016年 / 6卷

基金：

英国惠康基金; 美国国家卫生研究院;

关键词：

QUALITY-ASSURANCE;

D O I：

10.1038/srep20686

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

Standardization of immunophenotyping requires careful attention to reagents, sample handling, instrument setup, and data analysis, and is essential for successful cross-study and cross-center comparison of data. Experts developed five standardized, eight-color panels for identification of major immune cell subsets in peripheral blood. These were produced as pre-configured, lyophilized, reagents in 96-well plates. We present the results of a coordinated analysis of samples across nine laboratories using these panels with standardized operating procedures (SOPs). Manual gating was performed by each site and by a central site. Automated gating algorithms were developed and tested by the FlowCAP consortium. Centralized manual gating can reduce cross-center variability, and we sought to determine whether automated methods could streamline and standardize the analysis. Within-site variability was low in all experiments, but cross-site variability was lower when central analysis was performed in comparison with site-specific analysis. It was also lower for clearly defined cell subsets than those based on dim markers and for rare populations. Automated gating was able to match the performance of central manual analysis for all tested panels, exhibiting little to no bias and comparable variability. Standardized staining, data collection, and automated gating can increase power, reduce variability, and streamline analysis for immunophenotyping.

引用

页数：11

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