The Transglutaminase 2/TGM2 Assay Kit uses biotinylated T26 peptide (Biotin-pepT26) as the first substrate (amine-acceptor/acyl-donor) and an amine-donor/acyl-acceptor as a second substrate.
Samples suspected of containing TG2 are incubated with calcium, dithiothreitol (DTT) and Biotin-pepT26 in the wells of microtiter plates containing the amine substrate. In the presence of TG2, the gamma carboxamide of the glutaminyl residue of the biotin-pepT26 is incorporated into the amine substrate to form biotinylated isopetide bound.
The system is coupled to Streptavidin labelled peroxidase (SAv-HRP). SAv-HRP is revealed using H2O2 as HRP substrate and tetramethyl benzidine as electron acceptor (chromogen).
Measurement of tissue Transglutaminase (tTG/TG2) with low or no interference with other TGs isoforms.
Opr0036 : Recombinant human TG2 (positive control) (lyophilzed)
Microtiter strips coated with amine substrate (12x8 strip microtiter wells)
DTT (0.2 ml)
EDTA (Negative Control) (1 ml)
Enzyme Tracer (SAv-HRP) (50 ul)
Diluent Buffer 10X (10 ml)
HRP Substrate (12 ml)
Blocking Reagent (12 ml)
Wash Buffer 10X (30 ml)
The kit is shipped on ice. Upon arrival, the DTT (R1), the Reaction Buffer (R3) and Enzyme Tracer (R4) should be stored at -20C. All the other components of the kit should be kept at 4C. When stored properly, these stock solutions are stable for at least 24 months. OTHER SUPPLIES REQUIRED: - Recombinant human TG2 - TG2 inhibitor. This is recommended as negative control for screening of TG2 inhibitors.
Alternate Names for Transglutaminase 2/TGM2 Assay Kit (Colorimetric)
Transglutaminases (EC. 2.3.213, R- glutamyl-peptide amine g-glutamyl-transferase) are a family of calcium dependent enzymes which catalyse an acyl transferase reaction between the gamma-carboxamide group of peptide bound glutamine and various primary amines. In mammals, at least eight active transglutaminase (TGs) isoenzymes have been described so far. They are widely distributed in various organs, tissues and body fluids. Among them, transglutaminase type 2 (TG2) is distinguished from others TGs by its functional versatility and ubiquitous expression pattern in mammalian tissues. This isoenzyme is involved in a variety of roles including stabilization of intra and extracellular matrices and crosslinking of cell envelopes in apoptosis. It has also been associated with a large number of pathological conditions such as fibrosis, celiac disease, neurodegenerative disorders, inflammatory processes in sepsis, and in carcinogenesis of hepatocellular and ovarian carcinoma. This consideration has led to the development of several tests for either research or clinical purpose. Among the various strategies developed to measure TG activity, the most sensitive and accurate quantification methods, although non-discriminatory, have been the radioactive and fluorimetric assays. Those methods detect the incorporation of labeled molecules, such as [3H/14C]-putrescine or monodansyl-cadaverine respectively, into glutamyl substrates such as casein or synthetic peptides. Solid phase assays developed so far in similar principles have been compromise by a high background signal, low sensitivity compare to radiolabeling methods and the lack of the specificity. This test kit overcomes these problems and constitutes a highly sensitive and specific TG2 solid-phase microassay. The TG2-Covtest uses a biotinylated preferred first substrate of TG2 (biotin-pepT26) as amine-acceptor, and spermine as second substrate (amine-donor) of the enzyme. Samples suspected of containing TG2 are incubated with calcium, dithiothreitol (DTT) and biotin-pepT26 in the wells of microtiter plates to which spermine has been covalently coupled. In the presence of TG2, spermine is incorporated into the g carboxamide of the glutaminyl residue of biotin-pepT26 to form a biotin-pepT26- g-glutamyl spermine. Enzymatic reaction is determined by its interaction with Streptavidin labelled peroxidase (SAv-HRP). Following a wash step to remove any unbound enzyme reagent, a substrate solution for SAv-HRP containing H2O2 and tetramethyl benzidine as electron acceptor (chromogen) is added and color developed. The color intensity is directly proportional to the TG2 activity in the sample.
This product is for research use only and is not approved for use in humans or in clinical diagnosis. Kits are guaranteed for 6 months from date of receipt.
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FAQs for Transglutaminase 2/TGM2 Kit (NBP1-37008). (Showing 1 - 3 of 3 FAQs).
Does this kit contain recombinants using baculovirus expression system? If there are such recombinants, could you let us know the component name?
To my knowledge, this kit does not contain any recombinant proteins. See below for the component list: Microtiter strips with covalently bound spermine (12x 8-wells strips), DTT (0.2 mL), EDTA (Negative Control) (0.5 mL), Reaction Buffer (Biotin-pepT26/CaCl2) 2 vials (Lyophilized powder), Enzyme Tracer (SAv-HRP) (50 uL), Wash Buffer 10X (30 mL), Diluent Buffer 10X (10 mL), HRP Substrate (12 mL), Blocking Reagent (6 mL).
I would like to use the Transglutaminase 2 Kit to detect TGM2 activity in cell lysate from mouse tissue. The tissue protein is in RIPA buffer, I am not sure whether RIPA buffer is OK for using this Kit?
It is recommended to use lysate samples in Triton buffer with freezing/thawing cycles. Due to the denaturating properties of RIPA, the use of sample in RIPA buffer is not recommended. However, you may perform a cascade dilution of your samples up to 1/100 at least.
Does the Transglutaminase 2/TGM2 kit (NBP1-37008) measure total Transglutaminase 2/TGM2 or just active?
The Transglutaminase 2/TGM2 kit (NBP1-37008) only detects active TGM2.
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