High-Throughput Mass Cytometry Staining for Immunophenotyping Clinical Samples
Emily M Thrash, Katja Kleinsteuber, Emma S Hathaway, Matthew Nazzaro, Eric Haas, F Stephen Hodi, Mariano Severgnini, Emily M Thrash, Katja Kleinsteuber, Emma S Hathaway, Matthew Nazzaro, Eric Haas, F Stephen Hodi, Mariano Severgnini
Abstract
As mass cytometry (MC) is implemented in clinical settings, the need for robust, validated protocols that reduce batch effects between samples becomes increasingly important. Here, we present a streamlined MC workflow for high-throughput staining that generates reproducible data for up to 80 samples in a single experiment by combining reference sample spike-in and palladium-based mass-tag cell barcoding. Although labor intensive, this workflow decreases experimental variables and thus reduces technical error and mitigates batch effects.
Conflict of interest statement
K.K. is currently an employee of bluebird bio. F.S.H. serves as a consultant to Genentech, Bristol-Myers Squibb, Merck, Novartis, Amgen, Sanofi, Bayer, Pfizer, EMD Serono, Verastem, Aduro, Celldex, and Incyte.
© 2020 The Authors.
Figures
References
- Bagwell C.B., Inokuma M., Hunsberger B., Herbert D., Bray C., Hill B., Stelzer G., LI S., Kollipara A., Ornatsky O. Automated data cleanup for mass cytometry. Cytometry A. 2020;97:184–198.
- Holland M., Cunningham R., Seymour L., Kleinsteuber K., Cunningham A., Patel T., Manos M., Brennick R., Zhou J., Hodi F.S. Separation, banking, and quality control of peripheral blood mononuclear cells from whole blood of melanoma patients. Cell Tissue Bank. 2018;19:783–790.
- Kimball A.K., Oko L.M., Bullock B.L., Nemenoff R.A., van Dyk L.F., Clambey E.T. A beginner's guide to analyzing and visualizing mass cytometry data. J. Immunol. 2018;200:3–22.
- Kleinsteuber K., Corleis B., Rashidi N., Nchinda N., Lisanti A., Cho J.L., Medoff B.D., Kwon D., Walker B.D. Standardization and quality control for high-dimensional mass cytometry studies of human samples. Cytometry A. 2016;89:903–913.
- Lee B.H., Kelly G., Bradford S., Davila M., Guo X.V., Amir E.D., Thrash E.M., Solga M.D., Lannigan J., Sellers B. A modified injector and sample acquisition protocol can improve data quality and reduce inter-instrument variability of the helios mass cytometer. Cytometry A. 2019;95:1019–1030.
- Patel T., Cunningham A., Holland M., Daley J., Lazo S., Hodi F.S., Severgnini M. Development of an 8-color antibody panel for functional phenotyping of human CD8+ cytotoxic T cells from peripheral blood mononuclear cells. Cytotechnology. 2018;70:1–11.
- Schulz A.R., Baumgart S., Schulze J., Urbicht M., Grützkau A., Mei H.E. Stabilizing antibody cocktails for mass cytometry. Cytometry A. 2019;95:910–916.
Source: PubMed