Green Cathepsin B Assay
ICT’s Green Cathepsin B Assay allows researchers to detect and monitor intracellular cathepsin activity over time in vitro using live whole cells. This in vitro assay employs the quenched substrate Rhodamine 110-(RR)2, which fluoresces green upon cleavage by active cathepsin enzymes. Analyze samples using flow cytometry or fluorescence microscopy.
Green Cathepsin B Assay
Cathepsins are a group of protease enzymes typically found in the lysosome. Cathepsin B plays important roles in many cellular functions (like intracellular protein degradation, antigen processing, etc.). Elevated cathepsin activity is associated with many disease states such as cancer, Alzheimer’s disease, and so on.
The Green Cathepsin B Assay contains Rhodamine 110 Cathepsin B Substrate, which is a non-cytotoxic and cell membrane permeant substrate that fluoresces green upon cleavage by active cathepsins. To use Rhodamine 110-(RR)2, add the substrate directly to the cell culture medium, incubate, and analyze. Because the substrate is cell permeant, it easily penetrates the cell membrane and the membranes of the internal cellular organelles – no lysis or permeabilization steps are required. If active cathepsin enzymes are present, the quenched Rhodamine 110-(RR)2 substrate is cleaved, resulting in an increase in green fluorescence signal. Samples can be analyzed by flow cytometry or fluorescence microscopy. Hoechst 33342 is included in the kit to label nuclei.
R110-(RR)2
Cathepsin B
500 nm / 525 nm
Flow cytometry, Fluorescence microscope
Cell culture
R110-(RR)2 at ≤-20°C, other components at 2-8°C
Domestic: Overnight Delivery; International: Priority Shipping
United States
- Dilute 10X Cellular Assay Buffer 1:10 with diH2O.
- Prepare samples and controls in fresh cell culture medium or 1X Cellular Assay Buffer.
- Reconstitute R110-(RR)2 with 50 µL DMSO.
- Dilute R110-(RR)2 1:5 by adding 200 µL PBS.
- Add diluted R110-(RR)2 to each sample at 1:50 (e.g., add 10 µL of diluted R110-(RR)2 to 490 µL of sample).
- Incubate while protected from light.
- If desired, label with additional stains, such as Hoechst 33342, DAPI, or an antibody.
- Analyze with a flow cytometer or fluorescence microscope. R110-(RR)2 excites at 500 nm and emits at 525 nm.
Kit 9151: 25 Tests
1 vial of Rhodamine 110-(RR)2 substrate [R110-(RR)2], 25 Tests, #6691
1 vial of Hoechst 33342 Stain, 200 µg/mL (1 mL), #639
1 bottle of 10X Cellular Assay Buffer (15 mL), #6694
Kit Manual
Kit 9152: 100 Tests
4 vials of Rhodamine 110-(RR)2 substrate [R110-(RR)2], 25 Tests, #6691
1 vial of Hoechst 33342 Stain, 200 µg/mL (1 mL), #639
1 bottle of 10X Cellular Assay Buffer (60 mL), #6695
Kit Manual