Key features and details
- Assay type: Quantitative
- Detection method: Fluorescent
- Platform: Microplate reader
- Assay time: 1 hr
- Sample type: Inhibitor compounds
Product nameCathepsin D Inhibitor Assay Kit
See all Cathepsin D kits
Sample typeInhibitor compounds
Assay time1h 00m
Cathepsin D Inhibitor Screening Kit (ab126779) is a fluorescence-based assay that utilizes the preferred cathepsin D substrate sequence GKPILFFRLK(Dnp)-D-R-NH2) labeled with MCA. Cathepsin D will cleave the synthetic substrate to release the quenched fluorescent group MCA, which can then easily be measured using a fluorometer or fluorescence plate reader at Ex/Em = 328/460 nm. The relative efficacy of test inhibitors are compared to the positive control inhibitor, Pepstatin A (IC50 < 0.1 nM). The Cathepsin D assay is simple, straightforward, and can be adapted to 96-well plate assays and is suitable for high throughput screening (HTS).
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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 pro-apoptotic signals. Cathepsins are lysosomal enzymes that are also used as sensitive markers in various toxicological investigations.
Storage instructionsStore at -20°C. Please refer to protocols.
Components Identifier 100 tests Cathepsin D, human (Lyophilized) Green 1 vial CD Reaction Buffer NM 1 x 10ml CD Substrate Amber 1 x 0.2ml Pepstatin A (10 µM in DMSO) Blue 1 x 20µl
RelevanceCathepsin D is a normal lysosomal protease that is expressed in all cells. It is an aspartyl protease with a pH optimum in the range of 3-5, and contains two N linked oligosaccharides. Cathepsin D is synthesized as an inactive pro enzyme. Activation involves the proteolytic removal of the 43 amino acid profragment and an internal cleavage to generate the two chain form made up of 34 and 14 kDa subunits. Cathepsin D contains the mannose-6-phosphate lysosomal localization signal that targets the enzyme to the lysosomal compartment where it functions in the normal degradation of proteins. In certain tumor cells, Cathepsin D is abnormally processed and is secreted in its precursor form. Numerous clinical studies as well as in vitro evidence suggest that cathepsin D plays an important role in malignant transformation and may be a useful prognostic indicator for breast cancer and possibly Alzheimer's disease.
Cellular localizationLysosome. Melanosome. Note=Identified by mass spectrometry in melanosome fractions from stage I to stage IV.
- Lysosomal aspartyl protease
ab126779 has been referenced in 1 publication.
- Qiao S et al. The antimalarial amodiaquine causes autophagic-lysosomal and proliferative blockade sensitizing human melanoma cells to starvation- and chemotherapy-induced cell death. Autophagy 9:2087-102 (2013). PubMed: 24113242