Computational protocol: Analysis of cell surface and intranuclear markers on non-stimulated human PBMC using mass cytometry

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Protocol publication

[…] Cellular events were acquired on the Helios, which is available at the “Plateforme de Cytométrie de la Pitié-Salpêtrière”. Cells were acquired at a speed of 300 events per second, with a cell length threshold between 10 and 150 pushes. The 4 elements EQ beads were used to normalize files using the Matlab compiler software cell normalizer. Data files were obtained in the FCS file format and analysed using FlowJo software version 10 and Cytobank cloud based platform. Beads were gated out using the following gates: 140/142Ce, 151/153Eu, 165Ho and 175/176 Lu. CD45+ events were selected for the analysis of different cell populations. Cisplatin+ dead cells were gated out and we performed a viSNE (Stochastic Neighbor Embedding) analysis on CD45+ cells for auniform and non-biased separation of events. The settings used for the viSNE run were as follow: equal event sampling (20.000 events each), channels (CD19, CD4, IgD, CD16, CD14, CD8, CD56 and CD3), iterations (1000), perplexity (50) and theta (0.3). […]

Pipeline specifications

Software tools FlowJo, tSNE
Databases Cytobank
Application Flow cytometry
Organisms Homo sapiens
Diseases Autoimmune Diseases
Chemicals Palladium