PuSH - Publikationsserver des Helmholtz Zentrums München: Microfluidic proximity ligation assay for profiling signaling networks with single-cell resolution.

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Blazek, M.* ; Roth, G.* ; Zengerle, R.* ; Meier, M.

Microfluidic proximity ligation assay for profiling signaling networks with single-cell resolution.

Methods Mol. Biol. 1346, 169-184 (2015)

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Open Access Green möglich sobald Postprint bei der ZB eingereicht worden ist.

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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.

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