%0 Journal Article
%T A new branched proximity hybridization assay for the quantification of nanoscale protein¨Cprotein proximity
%A Jianying Yang
%A Marco Cavallari
%A Melanie Sieder
%A Michael Mitterer
%A Michael Reth
%A Shuangshuang Zheng
%J -
%D 2019
%R 10.1371/journal.pbio.3000569
%X Membrane proteins are organized in nanoscale compartments. Their reorganization plays a crucial role in receptor activation and cell signaling. To monitor the organization and reorganization of membrane proteins, we developed a new branched proximity hybridization assay (bPHA) allowing better quantification of the nanoscale protein¨Cprotein proximity. In this assay, oligo-coupled binding probes, such as aptamer, nanobody, and antibodies, are used to translate the proximity of target proteins to the proximity of oligos. The closely positioned oligos then serve as a template for a maximum of 400-fold branched DNA (bDNA) signal amplification. The amplified bPHA signal is recorded by flow cytometer, thus enabling proximity studies with high throughput, multiplexing, and single-cell resolution. To demonstrate the potential of the bPHA method, we measured the reorganization of the immunoglobulin M (IgM)- and immunoglobulin D (IgD)-class B cell antigen receptor (BCR) on the plasma membrane and the recruitment of spleen tyrosine kinase (Syk) to the BCR upon B lymphocyte activation
%K B cells
%K Flow cytometry
%K Cell staining
%K Green fluorescent protein
%K Cell hybridization
%K Antibodies
%K B cell receptors
%K Signal amplification
%U https://journals.plos.org/plosbiology/article?id=10.1371/journal.pbio.3000569