title: Transient partial permeabilization with saponin enables cellular barcoding prior to surface marker staining.
abstract: 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.
authors.affiliation: Baxter Laboratory for Stem Cell Biology, Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, California; Divisions of Hematology and Oncology, Department of Medicine, Stanford University School of Medicine, Stanford, California.
authors.forename: Gregory K
authors.initials: GK
authors.lastname: Behbehani
authors.forename: Colin
authors.initials: C
authors.lastname: Thom
authors.forename: Eli R
authors.initials: ER
authors.lastname: Zunder
authors.forename: Rachel
authors.initials: R
authors.lastname: Finck
authors.forename: Brice
authors.initials: B
authors.lastname: Gaudilliere
authors.forename: Gabriela K
authors.initials: GK
authors.lastname: Fragiadakis
authors.forename: Wendy J
authors.initials: WJ
authors.lastname: Fantl
authors.forename: Garry P
authors.initials: GP
authors.lastname: Nolan
mesh_terms: D003592:Cytophotometry / Q000379:methods*; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D019169:Jurkat Cells; D061848:Optical Imaging / Q000379:methods*; D012503:Saponins; D020298:U937 Cells
chemical_list: D012503:Saponins
keywords: fluorescent cellular barcoding; mass cytometry; mass-tag cellular barcoding; permeabilization; saponin