Transient partial permeabilization with saponin enables cellular barcoding prior to surface marker staining

Fluorescent cellular barcoding and mass‐tag cellular barcoding are cytometric methods that enable high sample throughput, minimize inter‐sample variation, and reduce reagent consumption. Previously employed barcoding protocols require that barcoding be performed after surface marker staining, compli...

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Bibliographic Details
Published in:Cytometry. Part A 2014-12, Vol.85 (12), p.1011-1019
Main Authors: Behbehani, Gregory K., Thom, Colin, Zunder, Eli R., Finck, Rachel, Gaudilliere, Brice, Fragiadakis, Gabriela K., Fantl, Wendy J., Nolan, Garry P.
Format: Article
Language:English
Subjects:
Cytophotometry - methods
fluorescent cellular barcoding
High-Throughput Screening Assays - methods
Humans
Jurkat Cells
mass cytometry
mass‐tag cellular barcoding
Optical Imaging - methods
permeabilization
saponin
Saponins
U937 Cells
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Summary:Fluorescent cellular barcoding and mass‐tag cellular barcoding are cytometric methods that enable high sample throughput, minimize inter‐sample variation, and reduce reagent consumption. Previously employed barcoding protocols require that barcoding be performed after surface marker staining, complicating combining the technique with measurement of alcohol‐sensitive surface epitopes. This report describes a method of barcoding fixed cells after a transient partial permeabilization with 0.02% saponin that results in efficient and consistent barcode staining with fluorescent or mass‐tagged reagents while preserving surface marker staining. This approach simplifies barcoding protocols and allows direct comparison of surface marker staining of multiple samples without concern for variations in the antibody cocktail volume, antigen‐antibody ratio, or machine sensitivity. Using this protocol, cellular barcoding can be used to reliably detect subtle differences in surface marker expression. © 2014 International Society for Advancement of Cytometry
ISSN:1552-4922
1552-4930
DOI:10.1002/cyto.a.22573