3D projection electrophoresis for single-cell immunoblotting

Author:

Grist Samantha M.ORCID,Mourdoukoutas Andoni P.,Herr Amy E.ORCID

Abstract

AbstractImmunoassays and mass spectrometry are powerful single-cell protein analysis tools; however, interfacing and throughput bottlenecks remain. Here, we introduce three-dimensional single-cell immunoblots to detect both cytosolic and nuclear proteins. The 3D microfluidic device is a photoactive polyacrylamide gel with a microwell array-patterned face (xy) for cell isolation and lysis. Single-cell lysate in each microwell is “electrophoretically projected” into the 3rd dimension (z-axis), separated by size, and photo-captured in the gel for immunoprobing and confocal/light-sheet imaging. Design and analysis are informed by the physics of 3D diffusion. Electrophoresis throughput is > 2.5 cells/s (70× faster than published serial sampling), with 25 immunoblots/mm2 device area (>10× increase over previous immunoblots). The 3D microdevice design synchronizes analyses of hundreds of cells, compared to status quo serial analyses that impart hours-long delay between the first and last cells. Here, we introduce projection electrophoresis to augment the heavily genomic and transcriptomic single-cell atlases with protein-level profiling.

Funder

National Science Foundation

U.S. Department of Health & Human Services | NIH | National Cancer Institute

Gouvernement du Canada | Natural Sciences and Engineering Research Council of Canada

Chan Zuckerberg Biohub

Publisher

Springer Science and Business Media LLC

Subject

General Physics and Astronomy,General Biochemistry, Genetics and Molecular Biology,General Chemistry

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