Chapter title |
Microfluidic Proximity Ligation Assay for Profiling Signaling Networks with Single-Cell Resolution
|
---|---|
Chapter number | 12 |
Book title |
Single Cell Protein Analysis
|
Published in |
Methods in molecular biology, January 2015
|
DOI | 10.1007/978-1-4939-2987-0_12 |
Pubmed ID | |
Book ISBNs |
978-1-4939-2986-3, 978-1-4939-2987-0
|
Authors |
Matthias Blazek, Günter Roth, Roland Zengerle, Matthias Meier |
Abstract |
The proximity ligation assay (PLA) is a technique that can be used to characterize proteins, protein-protein interactions, and protein modifications at the single-cell level. Image-based in situ detection of proteins using PLA is a quantitative method with a high degree of sensitivity and specificity. The miniaturization and parallelization of the PLA onto a microfluidic chip and concurrent use of an automated cell-culture system increase the throughput of this technology. Here, we describe the performance of PLA on a microfluidic chip. We provide protocols for on-chip cell culture, time-shifted cell stimulation and fixation, PLA implementation, and computational image analysis in order to achieve single-cell resolution. As a proof of concept, we studied the phosphorylation of Akt in response to stimulation with platelet-derived growth factor. |
Mendeley readers
Geographical breakdown
Country | Count | As % |
---|---|---|
Unknown | 12 | 100% |
Demographic breakdown
Readers by professional status | Count | As % |
---|---|---|
Researcher | 3 | 25% |
Student > Master | 2 | 17% |
Student > Bachelor | 1 | 8% |
Professor > Associate Professor | 1 | 8% |
Student > Postgraduate | 1 | 8% |
Other | 0 | 0% |
Unknown | 4 | 33% |
Readers by discipline | Count | As % |
---|---|---|
Biochemistry, Genetics and Molecular Biology | 4 | 33% |
Agricultural and Biological Sciences | 1 | 8% |
Immunology and Microbiology | 1 | 8% |
Medicine and Dentistry | 1 | 8% |
Unknown | 5 | 42% |