ArticleNature protocols2016
Single cell-resolution western blotting.
Article in Nature protocols, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 70 papers.
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Who cites it
70 citing papers in PubMed.
- Western Blot Analysis in Ewing Sarcoma.Methods in molecular biology (Clifton, N.J.) · 2027Article
- Single-Cell Protein Assays in Context: From 2D to 3D and In Situ Analysis.Annual review of analytical chemistry (Palo Alto, Calif.) · 2026Review
- Silk Fibroin Hydrogels: Cutting-edge Developments and Future Directions.Pharmaceutical nanotechnology · 2026Review
- Scaling Effects on Single-Cell Manipulation Using Magnetic Forces at Edge of Flat Plate With Elliptically Micro-Projection.Biotechnology and bioengineering · 2025Article
- Single-Cell Nucleus Extraction with Cellular Indexing.Advanced science (Weinheim, Baden-Wurttemberg, Germany) · 2025Article
- Protein Engineering and Drug Discovery: Importance, Methodologies, Challenges, and Prospects.Biomolecules · 2025Review
- Cytokine Storm Induction Linked to Multi-Organ Failure in Fatal Jellyfish Stings.Advanced science (Weinheim, Baden-Wurttemberg, Germany) · 2025Article
- Liquid biopsy of circulating tumor cells: From isolation, enrichment, and genome sequencing to clinical applications.Histology and histopathology · 2025Review
- Paired Analyses of Nuclear Protein Targets and Genomic DNA by Single-Cell Western Blot and Single-Cell PCR.Analytical chemistry · 2025Article
- Electromigration of Charged Analytes Through Immiscible Fluids in Multiphasic Electrophoresis.Electrophoresis · 2025Article
- Miniaturized protein profiling permits targeted signaling pathway analysis in individual circulating tumor cells to improve personalized treatment.Journal of translational medicine · 2024Article
- Article
- DropBlot: single-cell western blotting of chemically fixed cancer cells.Nature communications · 2024Article
- Band Degeneracy and Anisotropy Enhances Thermoelectric Performance from SbJournal of the American Chemical Society · 2024Article
- Electromigration of Charged Analytes Through Immiscible Fluids in Multiphasic Electrophoresis.bioRxiv : the preprint server for biology · 2024Article
- Pick-up single-cell proteomic analysis for quantifying up to 3000 proteins in a Mammalian cell.Nature communications · 2024Article
- Western blotting (immunoblotting): history, theory, uses, protocol and problems.BioTechniques · 2023Review
- Three-dimensional histological electrophoresis enables fast automatic distinguishment of cancer margins and lymph node metastases.Science advances · 2023Article
- Neural cell isolation from adult macaques for high-throughput analyses and neurosphere cultures.Nature protocols · 2023Review
- Presence of immunogenic alternatively spliced insulin gene product in human pancreatic delta cells.Diabetologia · 2023Article
10 more citing papers are in PubMed but not listed here.
Corrections and comments
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Authors and funding
6 authors.
Funding
Abstract
This protocol describes how to perform western blotting on individual cells to measure cell-to-cell variation in protein expression levels and protein state. Like conventional western blotting, single-cell western blotting (scWB) is particularly useful for protein targets that lack selective antibodies (e.g., isoforms) and in cases in which background signal from intact cells is confounding. scWB is performed on a microdevice that comprises an array of microwells molded in a thin layer of a polyacrylamide gel (PAG). The gel layer functions as both a molecular sieving matrix during PAGE and a blotting scaffold during immunoprobing. scWB involves five main stages: (i) gravity settling of cells into microwells; (ii) chemical lysis of cells in each microwell; (iii) PAGE of each single-cell lysate; (iv) exposure of the gel to UV light to blot (immobilize) proteins to the gel matrix; and (v) in-gel immunoprobing of immobilized proteins. Multiplexing can be achieved by probing with antibody cocktails and using antibody stripping/reprobing techniques, enabling detection of 10+ proteins in each cell. We also describe microdevice fabrication for both uniform and pore-gradient microgels. To extend in-gel immunoprobing to gels of small pore size, we describe an optional gel de-cross-linking protocol for more effective introduction of antibodies into the gel layer. Once the microdevice has been fabricated, the assay can be completed in 4-6 h by microfluidic novices and it generates high-selectivity, multiplexed data from single cells. The technique is relevant when direct measurement of proteins in single cells is needed, with applications spanning the fundamental biosciences to applied biomedicine.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.