廠牌: Immunochemistry 產地: 美國
FAM-FLICA® Caspase-8 Assay Kit
Size: 25 Tests
使用 FLICA Caspase-8 Assay Kit 檢測 caspase-8 活性。這種體外測定採用螢光抑製劑探針 FAM-LETD-FMK 來標記活細胞中的活性 caspase-8 酶。使用螢光顯微鏡、螢光盤式分析儀或流式細胞儀分析樣品。
Background 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 FAM FLICA Caspase-8 assay probe allows researchers to assess caspase-8 activation. The FLICA reagent FAM-LETD-FMK enters each cell and irreversibly binds to activated caspase-8. Because the FAM-LETD-FMK FLICA reagent becomes covalently coupled to the active enzyme, it is retained within the cell, while any unbound FAM-LETD-FMK FLICA reagent diffuses out of the cell and is washed away. The remaining green fluorescent signal is a direct measure of the active caspase-8 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 Propidium Iodide or Hoechst 33342 to detect necrosis or changes in nuclear morphology, respectively.
Reagent Name FAM-LETD-FMK
Excitation / Emission 488 nm / 530 nm
Method of Analysis Flow Cytometer, Fluorescence Microscope, Fluorescence Plate Reader
Sample Type Cell culture
Country of Origin United States
Prepare samples and controls
Dilute 10X Apoptosis Wash Buffer 1:10 with diH20.
Reconstitute FLICA with 50 μL DMSO.
Dilute FLICA 1:5 by adding 200 μL PBS.
Add diluted FLICA to each sample at 1:30 (e.g., add 10 μL to 290 μL of cultured cells).
Incubate approximately 1 hour.
Remove media and wash cells 3 times: add 1X Apoptosis Wash Buffer and spin cells.
If desired, label with additional stains, such as Hoechst, Propidium Iodide, 7-AAD, or an antibody.
If desired, fix cells.
Analyze with a fluorescence microscope, fluorescence plate reader, or flow cytometer. FAM-FLICA excites at 492 nm and emits at 520 nm.
****If working with adherent cells, please see the manual for additional protocols.
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