ArticleMolecular & cellular proteomics : MCP2013
Proximity ligation assay for high-content profiling of cell signaling pathways on a microfluidic chip.
Article in Molecular & cellular proteomics : MCP, 2013. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 17 papers.
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Who cites it
17 citing papers in PubMed, 29 citations in OpenAlex.
- Proximity Ligation Assay: From a Foundational Principle to a Versatile Platform for Molecular and Translational Research.Biomolecules · 2025Review
- Photosensitive Hydrogel with Temperature-Controlled Reversible Nano-Apertures for Single-Cell Protein Analysis.Advanced science (Weinheim, Baden-Wurttemberg, Germany) · 2024Article
- Engineering global and local signal generators for probing temporal and spatial cellular signaling dynamics.Frontiers in bioengineering and biotechnology · 2023Review
- A High-Throughput Single-Cell Assay on a Valve-Based Microfluidic Platform Applied to Protein Quantification, Immune Response Monitoring, and Drug Discovery.Methods in molecular biology (Clifton, N.J.) · 2023Article
- The Intriguing Landscape of Single-Cell Protein Analysis.Advanced science (Weinheim, Baden-Wurttemberg, Germany) · 2022Review
- A design and optimization of a high throughput valve based microfluidic device for single cell compartmentalization and analysis.Scientific reports · 2021Article
- Proximity ligation assay: an ultrasensitive method for protein quantification and its applications in pathogen detection.Applied microbiology and biotechnology · 2021Review
- Modular operation of microfluidic chips for highly parallelized cell culture and liquid dosing via a fluidic circuit board.Microsystems & nanoengineering · 2020Article
- Proximity Ligation Assays for In Situ Detection of Innate Immune Activation: Focus on In Vitro-Transcribed mRNA.Molecular therapy. Nucleic acids · 2019Article
- Integrating Immunology and Microfluidics for Single Immune Cell Analysis.Frontiers in immunology · 2018Review
- Scaling by shrinking: empowering single-cell 'omics' with microfluidic devices.Nature reviews. Genetics · 2017Review
- Efficient in situ detection of mRNAs using the Chlorella virus DNA ligase for padlock probe ligation.RNA (New York, N.Y.) · 2017Article
- In situ characterization of the mTORC1 during adipogenesis of human adult stem cells on chip.Proceedings of the National Academy of Sciences of the United States of America · 2016Article
- Digital microfluidic immunocytochemistry in single cells.Nature communications · 2015Article
- The nanoscale organization of the B lymphocyte membrane.Biochimica et biophysica acta · 2015Review
- Article
- Where is mTOR and what is it doing there?The Journal of cell biology · 2013Review
Corrections and comments
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Authors and funding
6 authors at 2 institutions in 2 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Here, we present the full integration of a proximity ligation assay (PLA) on a microfluidic chip for systematic cell signaling studies. PLA is an in situ technology for the detection of protein interaction, post-translational modification, concentration, and cellular location with single-molecule resolution. Analytical performance advances on chip are achieved, including full automation of the biochemical PLA steps, target multiplexing, and reduction of antibody consumption by 2 orders of magnitude relative to standard procedures. In combination with a microfluidic cell-culturing platform, this technology allows one to gain control over 128 cell culture microenvironments. We demonstrate the use of the combined cell culture and protein analytic assay on chip by characterizing the Akt signaling pathway upon PDGF stimulation. Signal transduction is detected by monitoring the phosphorylation states of Akt, GSK-3β, p70S6K, S6, Erk1/2, and mTOR and the cellular location of FoxO3a in parallel with the PLA. Single-cell PLA results revealed for Akt and direct targets of Akt a maximum activation time of 4 to 8 min upon PDGF stimulation. Activation times for phosphorylation events downward in the Akt signaling pathway including the phosphorylation of S6, p70S6K, and mTOR are delayed by 8 to 10 min or exhibit a response time of at least 1 h. Quantitative confirmation of the Akt phosphorylation signal was determined with the help of a mouse embryonic fibroblast cell line deficient for rictor. In sum, this work with a miniaturized PLA chip establishes a biotechnological tool for general cell signaling studies and their dynamics relevant for a broad range of biological inquiry.
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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.