ArticleAnalytical chemistry2016
Multiplexed Western Blotting Using Microchip Electrophoresis.
Article in Analytical chemistry, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 19 papers.
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Who cites it
19 citing papers in PubMed, 49 citations in OpenAlex.
- From Gold to Polymer: Latex Bead-Assisted CRISPR-Cas12a Platform for Next-Generation Protein Diagnostics.ACS measurement science au · 2026Article
- Comprehensive Optimization of Western Blotting.Gels (Basel, Switzerland) · 2023Article
- Multiplexed Fluorescence Plate Reader In Situ Protein Expression Assay in Apoptotic HepG2 Cells.International journal of molecular sciences · 2023Article
- Genome-wide identification of members of thePhysiology and molecular biology of plants : an international journal of functional plant biology · 2023Article
- Capillary electrophoresis Western blot using inkjet transfer to membrane.Journal of chromatography. A · 2022Article
- The Intriguing Landscape of Single-Cell Protein Analysis.Advanced science (Weinheim, Baden-Wurttemberg, Germany) · 2022Review
- Electrotransfer of Immunoprobes through Thin-Layer Polyacrylamide Gels.Analytical chemistry · 2022Article
- Summit: Automated Analysis of Arrayed Single-Cell Gel Electrophoresis.SLAS technology · 2021Article
- Microfluidics for Peptidomics, Proteomics, and Cell Analysis.Nanomaterials (Basel, Switzerland) · 2021Review
- Application of Microfluidics in Immunoassay: Recent Advancements.Journal of healthcare engineering · 2021Review
- 3D projection electrophoresis for single-cell immunoblotting.Nature communications · 2020Article
- Rapid electrotransfer probing for improved detection sensitivity in in-gel immunoassays.Analytical methods : advancing methods and applications · 2020Article
- Fast Immunoassay for Microfluidic Western Blotting by Direct Deposition of Reagents onto Capture Membrane.Analytical methods : advancing methods and applications · 2020Article
- Antibody validation for Western blot: By the user, for the user.The Journal of biological chemistry · 2020Article
- Controlling Dispersion during Single-Cell Polyacrylamide-Gel Electrophoresis in Open Microfluidic Devices.Analytical chemistry · 2018Article
- Review
- Protein purification and analysis: next generation Western blotting techniques.Expert review of proteomics · 2017Review
- Review
- Recent Advances in Microscale Western Blotting.Analytical methods : advancing methods and applications · 2016Article
Corrections and comments
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Authors and funding
8 authors at 2 institutions in 1 country.
Funding
Abstract
Western blotting is a commonly used protein assay that combines the selectivity of electrophoretic separation and immunoassay. The technique is limited by long time, manual operation with mediocre reproducibility, and large sample consumption, typically 10-20 μg per assay. Western blots are also usually used to measure only one protein per assay with an additional housekeeping protein for normalization. Measurement of multiple proteins is possible; however, it requires stripping membranes of antibody and then reprobing with a second antibody. Miniaturized alternatives to Western blot based on microfluidic or capillary electrophoresis have been developed that enable higher-throughput, automation, and greater mass sensitivity. In one approach, proteins are separated by electrophoresis on a microchip that is dragged along a polyvinylidene fluoride membrane so that as proteins exit the chip they are captured on the membrane for immunoassay. In this work, we improve this method to allow multiplexed protein detection. Multiple injections made from the same sample can be deposited in separate tracks so that each is probed with a different antibody. To further enhance multiplexing capability, the electrophoresis channel dimensions were optimized for resolution while keeping separation and blotting times to less than 8 min. Using a 15 μm deep × 50 μm wide × 8.6 cm long channel, it is possible to achieve baseline resolution of proteins that differ by 5% in molecular weight, e.g., ERK1 (44 kDa) from ERK2 (42 kDa). This resolution allows similar proteins detected by cross-reactive antibodies in a single track. We demonstrate detection of 11 proteins from 9 injections from a single Jurkat cell lysate sample consisting of 400 ng of total protein using this procedure. Thus, multiplexed Western blots are possible without cumbersome stripping and reprobing steps.
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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.