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Within the field of molecular biology, a protein-fragment complementation assay, or PCA, is a method for the identification and quantification of protein–protein interactions. In the PCA, the proteins of interest ("bait" and "prey") are each covalently linked to fragments of a third protein (e.g. DHFR, which acts as a "reporter").
To perform immunofluorescence staining, a fluorophore must be conjugated (“tagged”) to an antibody. Staining procedures can be applied to both retained intracellular expressed antibodies, or to cell surface antigens on living cells.
In-vivo crosslinking of protein complexes using photo-reactive amino acid analogs was introduced in 2005 by researchers from the Max Planck Institute [5] In this method, cells are grown with photoreactive diazirine analogs to leucine and methionine, which are incorporated into proteins.
The sensitivity and high signal intensity of this luciferase molecule proves advantageous in many reporter studies. Some of the benefits of using a secreted reporter molecule like MetLuc is its no-lysis protocol that allows one to be able to conduct live cell assays and multiple assays on the same cell. [8]
Luciferase systems are widely used in genetic engineering as reporter genes, each producing a different color by fluorescence, [76] [77] and for biomedical research using bioluminescence imaging. [ 78 ] [ 79 ] [ 80 ] For example, the firefly luciferase gene was used as early as 1986 for research using transgenic tobacco plants. [ 81 ]
Immunoprecipitation of intact protein complexes (i.e. antigen along with any proteins or ligands that are bound to it) is known as co-immunoprecipitation (Co-IP). Co-IP works by selecting an antibody that targets a known protein that is believed to be a member of a larger complex of proteins.
Renilla-luciferin 2-monooxygenase, Renilla luciferase, or RLuc, is a bioluminescent enzyme found in Renilla reniformis, belonging to a group of coelenterazine luciferases.Of this group of enzymes, the luciferase from Renilla reniformis has been the most extensively studied, and due to its bioluminescence requiring only molecular oxygen, has a wide range of applications, with uses as a reporter ...
A bioassay is an analytical method to determine the potency or effect of a substance by its effect on living animals or plants (in vivo), or on living cells or tissues (in vitro). [1] [2] A bioassay can be either quantal or quantitative, direct or indirect. [3] If the measured response is binary, the assay is quantal; if not, it is quantitative ...