期刊
BIO-PROTOCOL
卷 7, 期 21, 页码 -出版社
BIO-PROTOCOL
DOI: 10.21769/BioProtoc.2602
关键词
Proximal ligation assay; Protein-protein interaction; Toll-like receptors; Fluorescence microscopy; Bronchial epithelium; Macrophages; J774; Duolink
类别
资金
- NIH [R21HL125021]
In this protocol, we describe proximal ligation assay (PLA), an antibody-based detection method for protein-protein interaction. This method relies on specific binding of individual primary antibodies to the two putative interacting proteins. The primary antibodies need to have different hosts. The secondary antibodies against the two hosts have complementary oligonucleotide moieties attached to them. If the two antigens are in close proximity (presumably interacting with each other), the complementary oligonucleotides can anneal and fluorescent nucleotides can be incorporated in a single DNA polymerization step. Under a microscope, these reactions appear as punctate fluorescent spots, indicating successful PLA reaction and suggesting protein-protein interaction between the two antigens.
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