Send to

Choose Destination
J Immunol. 2017 Feb 15;198(4):1748-1758. doi: 10.4049/jimmunol.1601750. Epub 2017 Jan 9.

An Integrated Workflow To Assess Technical and Biological Variability of Cell Population Frequencies in Human Peripheral Blood by Flow Cytometry.

Author information

La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037;
J. Craig Venter Institute, La Jolla, CA 92037.
La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037.
Division of Infectious Diseases, University of California, San Diego, La Jolla, CA 92093.
Johns Hopkins University Bloomberg School of Public Health, Baltimore, MD 21205.
Universidad Peruana Caytano Hereida, Lima 15102, Peru.
Department of Virology, Tohoku University, Sendai 9808575, Japan.
Genetech Research Center, Colombo 00800, Sri Lanka; and.
Department of Pathology, University of California, San Diego, La Jolla, CA 92093.


In the context of large-scale human system immunology studies, controlling for technical and biological variability is crucial to ensure that experimental data support research conclusions. In this study, we report on a universal workflow to evaluate both technical and biological variation in multiparameter flow cytometry, applied to the development of a 10-color panel to identify all major cell populations and T cell subsets in cryopreserved PBMC. Replicate runs from a control donation and comparison of different gating strategies assessed the technical variability associated with each cell population and permitted the calculation of a quality control score. Applying our panel to a large collection of PBMC samples, we found that most cell populations showed low intraindividual variability over time. In contrast, certain subpopulations such as CD56 T cells and Temra CD4 T cells were associated with high interindividual variability. Age but not gender had a significant effect on the frequency of several populations, with a drastic decrease in naive T cells observed in older donors. Ethnicity also influenced a significant proportion of immune cell population frequencies, emphasizing the need to account for these covariates in immune profiling studies. We also exemplify the usefulness of our workflow by identifying a novel cell-subset signature of latent tuberculosis infection. Thus, our study provides a universal workflow to establish and evaluate any flow cytometry panel in systems immunology studies.

[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for HighWire Icon for PubMed Central
Loading ...
Support Center