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Proteasome Activity Assay Kit (ab107921) takes advantage of the chymotrypsin-like activity, utilizing an AMC-tagged peptide substrate (Proteasome Substrate (Succ-LLVY-AMC in DMSO), which releases free, highly fluorescent AMC in the presence of proteolytic activity. The kit also includes a positive control (Jurkat Cell lysate with significant proteasome activity) and a specific proteasome inhibitor MG-132 which suppresses all proteolytic activity due to proteasomes. This permits differentiation of proteasome activity from other protease activity which may be present in samples.
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Proteasomes are very large (20S, 26S) protein assemblies found in both the nucleus and cytoplasm of all eucaryotes (and in some procaryotes). They are responsible for the degradation and recycling of proteins which have been previously tagged with ubiquitin. Such tagged proteins are degraded into peptides approximately 7-8 amino acids long which are subsequently further degraded. The 20S assembly is the functional protease structure with chymotrypsin-like, trypsin-like and caspase-like protease activities
|AMC Standard (1mM in DMSO)||Yellow||1 x 100µl|
|Positive Control (lyophilized)||Green||1 vial|
|Proeasome Substrate (in DMSO)||Red||1 x 100µl|
|Proteasome Assay Buffer||WM||1 x 25ml|
|Proteasome Inhibitor (in DMSO)||Blue||1 x 100µl|
Standard curve: mean of duplicates (+/- SD) with background reads subtracted
Proteasome Activity measured in Jurkat lysate
Proteasome Activity measured in HeLa lysate
Proteasome Activity measured in cell lysates showing activity (mU) per 1 mln cells.
Samples with the concentration of 3.3e6 cells/mL were used. 50 μl of each sample was used.
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