title: Automated flow cytometric analysis across large numbers of samples and cell types.
abstract: Multi-parametric flow cytometry is a key technology for characterization of immune cell phenotypes. However, robust high-dimensional post-analytic strategies for automated data analysis in large numbers of donors are still lacking. Here, we report a computational pipeline, called FlowGM, which minimizes operator input, is insensitive to compensation settings, and can be adapted to different analytic panels. A Gaussian Mixture Model (GMM)-based approach was utilized for initial clustering, with the number of clusters determined using Bayesian Information Criterion. Meta-clustering in a reference donor permitted automated identification of 24 cell types across four panels. Cluster labels were integrated into FCS files, thus permitting comparisons to manual gating. Cell numbers and coefficient of variation (CV) were similar between FlowGM and conventional gating for lymphocyte populations, but notably FlowGM provided improved discrimination of "hard-to-gate" monocyte and dendritic cell (DC) subsets. FlowGM thus provides rapid high-dimensional analysis of cell phenotypes and is amenable to cohort studies.
authors.affiliation: Systems Biology Lab, Institut Pasteur, Paris, France; Laboratory of Analysis Geometry and Modeling, Department of Mathematics, University of Cergy-Pontoise, Ile de France, France.
authors.forename: Xiaoyi
authors.initials: X
authors.lastname: Chen
authors.affiliation: Center for Human Immunology, Institut Pasteur, Paris France.
authors.forename: Milena
authors.initials: M
authors.lastname: Hasan
authors.affiliation: Center for Human Immunology, Institut Pasteur, Paris France.
authors.forename: Valentina
authors.initials: V
authors.lastname: Libri
authors.affiliation: Center for Human Immunology, Institut Pasteur, Paris France; INSERM U818, France; Laboratory of Dendritic Cell Immunobiology, Department of Immunology, Institut Pasteur, Paris France.
authors.forename: Alejandra
authors.initials: A
authors.lastname: Urrutia
authors.affiliation: Center for Human Immunology, Institut Pasteur, Paris France.
authors.forename: Benoît
authors.initials: B
authors.lastname: Beitz
authors.affiliation: Center for Bioinformatics, Institut Pasteur, Paris France.
authors.forename: Vincent
authors.initials: V
authors.lastname: Rouilly
authors.affiliation: Center for Human Immunology, Institut Pasteur, Paris France; INSERM U818, France; Laboratory of Dendritic Cell Immunobiology, Department of Immunology, Institut Pasteur, Paris France.
authors.forename: Darragh
authors.initials: D
authors.lastname: Duffy
authors.affiliation: Laboratory of Immunoregulation, Department of Immunology, Institut Pasteur, Paris France.
authors.forename: Étienne
authors.initials: É
authors.lastname: Patin
authors.affiliation: University of Cergy-Pontoise, France; CLMA, ENS-Cachan, France.
authors.forename: Bernard
authors.initials: B
authors.lastname: Chalmond
authors.affiliation: Center for Human Immunology, Institut Pasteur, Paris France; Laboratory of Immunoregulation, Department of Immunology, Institut Pasteur, Paris France.
authors.forename: Lars
authors.initials: L
authors.lastname: Rogge
authors.affiliation: Unit of Human Evolutionary Genetics, Department of Genomes & Genetics, Institut Pasteur, Paris, France; CNRS URA3012, France.
authors.forename: Lluis
authors.initials: L
authors.lastname: Quintana-Murci
authors.affiliation: Center for Human Immunology, Institut Pasteur, Paris France; INSERM U818, France; Laboratory of Dendritic Cell Immunobiology, Department of Immunology, Institut Pasteur, Paris France; INSERM UMS20, France. Electronic address: albertm@pasteur.fr.
authors.forename: Matthew L
authors.initials: ML
authors.lastname: Albert
authors.affiliation: Systems Biology Lab, Institut Pasteur, Paris, France. Electronic address: benno@pasteur.fr.
authors.forename: Benno
authors.initials: B
authors.lastname: Schwikowski
mesh_terms: D000465:Algorithms; D057205:Automation, Laboratory / Q000379:methods*; D001402:B-Lymphocytes; D001499:Bayes Theorem; D016000:Cluster Analysis; D003713:Dendritic Cells; D005434:Flow Cytometry / Q000379:methods*; D006801:Humans; D007694:Killer Cells, Natural; D009000:Monocytes; D009504:Neutrophils; D012015:Reference Standards; D012984:Software; D013223:Statistics as Topic; D016176:T-Lymphocyte Subsets; D013601:T-Lymphocytes
keywords: Algorithms;; Automation;; Flow cytometry;; Multidimensional analysis;; Population-based cohort;; Standardization;