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Abcam's Cathepsin D Activity Assay Kit is a fluorescence-based assay that utilizes the preferred cathepsin-D substrate sequence GKPILFFRLK(Dnp)-D-R-NH2) labeled with MCA. Cell lysates or other samples that contain cathepsin-D will cleave the synthetic substrate to release fluorescence, which can then easily be quantified using a fluorometer or fluorescence plate reader at Ex/Em = 328/460 nm.
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Apoptosis can be mediated by mechanisms other than the traditional caspase-mediated cleavage cascade. There is growing recognition that alternative proteolytic enzymes such as the lysosomal cathepsin proteases may initiate or propagate proapoptotic signals. Cathepsins are lysosomal enzymes that are also used as sensitive markers in various toxicological investigations.
|CD Cell Lysis Buffer||WM||1 x 25ml|
|CD Reaction Buffer||NM||1 x 5ml|
|CD Substrate (1mM)||Brown||1 x 200µl|
Enzyme activity of Cathepsin B and Cathepsin D was determined using Cathepsin B/D activity assay kit (ab65300 and ab65302). Neuro2a cell lysates were centrifuged at 10,000 xg for 10 minutes at 4ºC and supernatant was used for the assay. Cathepsin activity was performed at the 24 and 48 hours time points by cleavage of the fluorescence peptide substrate [DnP-DR-MCA, GKPILFFRLK(DnP)-DR substrate peptide labeled with MCA]. Data represent the mean ± SD (*p<0.05, **p<0.001).
Cathepsin D levels were measured in standard control samples from ab119586; background signal subtracted (duplicates +/- SD).
Cathepsin D measured in mouse tissue lysates (mg of extracted protein), background signal subtracted (duplicates +/- SD).
Cathepsin D measured in cell lysates, background signal subtracted (duplicates +/- SD).
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