title: Single-Cell Western Blotting.
abstract: Little headway has been made in single cell protein analysis, aside from tools that rely solely on antibody-probe based detection (i.e., flow cytometry, immunocytochemistry), which are limited by low specificity and multiplexing capabilities. To address these protein analysis gaps, we have introduced a single-cell western blot (scWestern). The protein assay is capable of highly specific analysis by coupling antibody-based detection with a polyacrylamide gel electrophoresis (PAGE) protein separation. Cells are settled via gravity into polyacrylamide (PA) microwells, chemically lysed in the wells, and then subjected to PAGE through the walls of the microwells and into the surrounding PA gel. Over a thousand single-cell separations are performed simultaneously, and multiple protein targets of interest are investigated. After PAGE separation, photo-immobilization of all proteins to the gel allows for antibody probing and lends to the archival quality of the scWestern assay where new proteins targets can be investigated months after the initial separations are performed.
authors.affiliation: Department of Bioengineering, University of California Berkeley, 306 Stanley Hall MC 1762, Berkeley, CA, 94720-1762, USA.
authors.forename: Elly
authors.initials: E
authors.lastname: Sinkala
authors.affiliation: Department of Bioengineering, University of California Berkeley, 306 Stanley Hall MC 1762, Berkeley, CA, 94720-1762, USA. aeh@berkeley.edu.
authors.forename: Amy E
authors.initials: AE
authors.lastname: Herr
mesh_terms: D000818:Animals; D015153:Blotting, Western / Q000295:instrumentation / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel / Q000295:instrumentation / Q000379:methods*; D004867:Equipment Design; D006801:Humans; D056656:Lab-On-A-Chip Devices; D011506:Proteins / Q000032:analysis*; D059010:Single-Cell Analysis / Q000295:instrumentation / Q000379:methods*
chemical_list: D011506:Proteins
keywords: Immunoassay; Immunoblot; Lab-on-a-chip; Proteomics; Single cell