Proximity Ligation Assay

from Wikipedia, the free encyclopedia

The Proximity Ligation Assay ( in situ PLA ) is a biochemical method for the detection of protein-protein interactions . The PLA is a combination of an ELISA with a signal amplification through a polymerase chain reaction and is therefore related to an immuno-polymerase chain reaction .

principle

Antibodies of different species are required against both suspected interaction partners. After two interaction partners bind to one another, detection is carried out using different immunoconjugates . The immunoconjugates carry different oligonucleotides which , due to the spatial proximity, together form a ring after adding two adapter oligonucleotides, which after ligation is replicated in a polymerase chain reaction by rolling circle replication . Interactions of a few molecules can thus be detected, but specific antibodies must be present for both interaction partners. This assumes the knowledge of both partners, which allows the method to be used to confirm the results of other methods of determining protein-protein interactions. With a fluorescence- labeled hybridization probe , the interaction in cells and tissue sections can be detected with a fluorescence microscope due to the fluorescent staining in comparison to the negative controls .

Individual evidence

  1. ↑ O. Söderberg, M. Gullberg, M. Jarvius, K. Ridderstråle, KJ Leuchowius, J. Jarvius, K. Wester, P. Hydbring, F. Bahram, LG Larsson, U. Landegren: Direct observation of individual endogenous protein complexes in situ by proximity ligation. In: Nature methods. Volume 3, Number 12, December 2006, ISSN  1548-7091 , pp. 995-1000, doi : 10.1038 / nmeth947 . PMID 17072308 .
  2. ↑ M. Jarvius, J. Paulsson, I. Weibrecht, KJ Leuchowius, AC Andersson, C. Wählby, M. Gullberg, J. Botling, T. Sjöblom, B. Markova, A. Ostman, U. Landegren, O. Söderberg : In situ detection of phosphorylated platelet-derived growth factor receptor beta using a generalized proximity ligation method. In: Molecular & cellular proteomics: MCP. Volume 6, Number 9, September 2007, ISSN  1535-9476 , pp. 1500-1509, doi : 10.1074 / mcp.M700166-MCP200 . PMID 17565975 .