SR-FLICA® Caspase-9 Assay Kit

$195.00$561.00

Detect caspase-9 activity with the SR-FLICA Caspase-9 Assay Kit. This in vitro assay employs the fluorescent inhibitor probe SR-LEHD-FMK to label active caspase-9 enzyme in living cells. Analyze samples using fluorescence microscopy, a fluorescence plate reader, or flow cytometry.

Catalog # Size Quantity Price
960 25 Tests $195.00
961 100 Tests $561.00
Catalog #: 960, 961 Categories: , ,

Caspases play important roles in apoptosis and inflammation. ICT’s FLICA assay kits are used by researchers seeking to quantitate apoptosis via caspase activity in cultured cells and tissues.

The FLICA reagent SR-LEHD-FMK enters each cell and irreversibly binds to activated caspase-9. Because the SR-LEHD-FMK FLICA reagent becomes covalently coupled to the active enzyme, it is retained within the cell, while any unbound SR-LEHD-FMK FLICA reagent diffuses out of the cell and is washed away. The remaining red fluorescent signal is a direct measure of the active caspase-9 enzyme activity present in the cell at the time the reagent was added. Cells that contain the bound FLICA can be analyzed by a fluorescence plate reader, fluorescence microscopy, or flow cytometry. Cells labeled with the FLICA reagent may be read immediately or preserved for 16 hours using the fixative included in the kit. Unfixed samples may also be analyzed with Hoechst 33342 to detect changes in nuclear morphology.

1. Prepare samples and controls.
2. Dilute 10X Apoptosis Wash Buffer 1:10 with diH20.
3. Reconstitute FLICA with 50 μL DMSO.
4. Dilute FLICA 1:5 by adding 200 μL PBS.
5. Add diluted FLICA to each sample at 1:30 – 1:60. For example, to stain at 1:30, add 10 μL to 290 μL of cultured cells. To stain at 1:60, add 5 μL to 295 μL of cultured cells.
6. Incubate approximately 1 hour.
7. Remove media and wash cells 3 times: add 1X Apoptosis Wash Buff er and spin cells.
8. If desired, label with additional stains, such as Hoechst, 7-AAD, or an antibody.
9. If desired, fix or embed cells.
10. Analyze with a fluorescence microscope, fluorescence plate reader, or flow cytometer. SR-FLICA excites at 550-580 nm and emits at 590-600 nm.

Jurkat cells were treated with 1 µM staurosporine to induce caspase-9 activity (left), or a negative control (right) for 3 hours, washed twice, then incubated with ICT’s red caspase-9 inhibitor probe, SR-LEHD-FMK, for 1 hour and examined under a fluorescence microscope. DIC images were also taken of all cells. Both induced images reveal several experimental cells which fluoresce red, therefore they have some degree of caspase-9 activity. The non-induced DIC image reveals many control cells, however the corresponding fluorescence image is dark; none of these cells have active caspase-9 (Dr. Brian W. Lee, ICT).
srlehddata

• Reagent name: SR-LEHD-FMK
• Target: Caspase-9
• Excitation/Emission: 550-580 nm / 590-600 nm
• Method of Analysis: Flow Cytometer, Fluorescence Microscope, Fluorescence Plate Reader
• Types of Samples: Cell culture
• Storage: 2-8°C
• Shipping: Ships overnight (domestic), International Priority Shipping

Kit 960: 25 Tests
• FLICA Caspase-9 Reagent (SR-LEHD-FMK), 1 vial, #6145
• 10X Apoptosis Wash Buffer, 15 mL, #635
• Fixative, 6 mL, #636
• Hoechst 33342, 1 mL, #639
• Kit Manual

Kit 961: 100 Tests
• FLICA Caspase-9 Reagent (SR-LEHD-FMK), 4 vials, #6145
• 10X Apoptosis Wash Buffer, 60 mL, #634
• Fixative, 6 mL, #636
• Hoechst 33342, 1 mL, #639
• Kit Manual

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