Mass cytometry immunostaining protocol for multiplexing clinical samples

Ramy Gadalla; Giselle M Boukhaled; David G Brooks; Ben X Wang

doi:10.1016/j.xpro.2022.101643

Mass cytometry immunostaining protocol for multiplexing clinical samples

STAR Protoc. 2022 Aug 22;3(3):101643. doi: 10.1016/j.xpro.2022.101643. eCollection 2022 Sep 16.

Authors

Ramy Gadalla¹, Giselle M Boukhaled², David G Brooks², Ben X Wang³

Affiliations

¹ Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2M9, Canada. Electronic address: ramy.gadalla@uhnresearch.ca.
² Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2M9, Canada; Department of Immunology, University of Toronto, Toronto, ON M5S 1A8, Canada.
³ Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2M9, Canada. Electronic address: ben.wang@uhnresearch.ca.

Abstract

This is a cytometry by time-of-flight (CyTOF) staining protocol for hematopoietic-derived cells, that leverages live-cell barcoding using receptor-type tyrosine-protein phosphatase C (CD45) antibodies conjugated to metal isotopes in combination with DNA-based palladium barcoding to multiplex up to 40 samples. In this protocol, DNA-based barcoding is performed before surface and intracellular immunostaining, which reduces the batch effects that result from day-to-day variations in staining and instrument sensitivity. This protocol also reduces antibody consumption and eliminates the need for repeated instrument adjustment.

Keywords: Flow Cytometry/Mass Cytometry; Immunology.

Publication types

Research Support, Non-U.S. Gov't
Research Support, N.I.H., Extramural

MeSH terms

Antibodies*
Flow Cytometry / methods
Isotopes*
Palladium
Staining and Labeling

Substances

Antibodies
Isotopes
Palladium

Grants and funding

FDN148386 /CIHR/Canada