Pyroptosis/Caspase-1 Assay Assay Kit
Pyroptosis/Caspase-1 Assay, Far Red
Applications
Flow Cytometer, Fluorescence Microscope
Synonyms
Pyroptosis/Caspase-1 Assay; Far Red; Far Red;Caspase-1; Pyroptosis/Caspase-1 Assay assay kit
Applicable Applications for Pyroptosis/Caspase-1 Assay assay kit
Flow Cytometer, Fluorescence Microscope
Application Notes
1. Prepare samples and controls.
2. Dilute 10X Cellular Assay Buffer 1:10 with diH2O.
3. Reconstitute FLICA with 50 uL DMSO.
4. Dilute FLICA 1:5 by adding 200 uL PBS.
5. Add diluted FLICA to each sample at 1:30-1:60 (e.g., spike at 1:30 by adding 10 uL to 290 uL sample).
6. Incubate approximately 1 hour.
7. Remove media and wash cells 3 times: add 1X Cellular Wash Buffer and spin cells.
8. Resuspend cell pellet in 1X Cellular Wash Buffer.
9. If desired, label with additional stains, such as Hoechst 33342, DAPI, or an antibody.
10. If desired, fix cells.
11. Analyze with a fluorescence microscope or flow cytometer. FLICA 660 is excited at 660 nm and emits at 680-690 nm.
2. Dilute 10X Cellular Assay Buffer 1:10 with diH2O.
3. Reconstitute FLICA with 50 uL DMSO.
4. Dilute FLICA 1:5 by adding 200 uL PBS.
5. Add diluted FLICA to each sample at 1:30-1:60 (e.g., spike at 1:30 by adding 10 uL to 290 uL sample).
6. Incubate approximately 1 hour.
7. Remove media and wash cells 3 times: add 1X Cellular Wash Buffer and spin cells.
8. Resuspend cell pellet in 1X Cellular Wash Buffer.
9. If desired, label with additional stains, such as Hoechst 33342, DAPI, or an antibody.
10. If desired, fix cells.
11. Analyze with a fluorescence microscope or flow cytometer. FLICA 660 is excited at 660 nm and emits at 680-690 nm.
Sample
Cell culture
Reagent Name
660-YVAD-FMK
Excitation/Emission
660 nm / 690 nm
Preparation and Storage
Store at 2-8 degree C.
Related Product Information for Pyroptosis/Caspase-1 Assay assay kit
Description: Pyroptosis/Caspase-1 Assay, Far Red utilizes our popular FLICA technology to measure caspase-1 activation. Using this kit, researchers can easily detect pyroptotic cells in their whole cell samples in vitro. This complete kit includes Nigericin for the simple and reliable generation of a positive control. Analyze the fluorescence signal using flow cytometry or fluorescence microscopy.
Background: Inflammatory caspases, such as caspase-1, play a critical role in the mechanism to trigger the lytic programmed cell death known as pyroptosis. Pyroptosis/Caspase-1 Assay, Far Red can be used by researchers to assess caspase-1 activity in pyroptotic cells.
This kit utilizes the far red, cell permeant FLICA reagent, 660-YVAD-FMK, for the in vitro detection of caspase-1 in whole living cells. 660-YVAD-FMK enters each cell and irreversibly binds to activated caspase-1. Because 660-YVAD-FMK becomes covalently coupled to the active enzyme, it is retained within the cell, while any unbound reagent diffuses out of the cell and is washed away. The remaining far red fluorescent signal is a direct measure of the active caspase-1 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 microscope or flow cytometer. Cells labeled with the FLICA reagent may be read immediately, or preserved for up to 16 hours using Fixative (included in the kit). Unfixed samples may also be counter-stained with Hoechst 33342 (included in the kit) to label cell nuclei. This complete kit also includes Nigericin as a convenient option for generating a positive control. Nigericin induces a net decrease in intracellular levels of potassium, crucial for activation of caspase-1. In pyroptosis experiments, it has been shown to generate robust caspase-1 activation in a variety of cell types.
Background: Inflammatory caspases, such as caspase-1, play a critical role in the mechanism to trigger the lytic programmed cell death known as pyroptosis. Pyroptosis/Caspase-1 Assay, Far Red can be used by researchers to assess caspase-1 activity in pyroptotic cells.
This kit utilizes the far red, cell permeant FLICA reagent, 660-YVAD-FMK, for the in vitro detection of caspase-1 in whole living cells. 660-YVAD-FMK enters each cell and irreversibly binds to activated caspase-1. Because 660-YVAD-FMK becomes covalently coupled to the active enzyme, it is retained within the cell, while any unbound reagent diffuses out of the cell and is washed away. The remaining far red fluorescent signal is a direct measure of the active caspase-1 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 microscope or flow cytometer. Cells labeled with the FLICA reagent may be read immediately, or preserved for up to 16 hours using Fixative (included in the kit). Unfixed samples may also be counter-stained with Hoechst 33342 (included in the kit) to label cell nuclei. This complete kit also includes Nigericin as a convenient option for generating a positive control. Nigericin induces a net decrease in intracellular levels of potassium, crucial for activation of caspase-1. In pyroptosis experiments, it has been shown to generate robust caspase-1 activation in a variety of cell types.
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Product Notes
The Pyroptosis/Caspase-1 Assay (Catalog #AAA258379) is an Assay Kit and is intended for research purposes only. The product is available for immediate purchase. AAA Biotech's Pyroptosis/Caspase-1 Assay can be used in a range of immunoassay formats including, but not limited to, Flow Cytometer, Fluorescence Microscope. 1. Prepare samples and controls. 2. Dilute 10X Cellular Assay Buffer 1:10 with diH2O. 3. Reconstitute FLICA with 50 uL DMSO. 4. Dilute FLICA 1:5 by adding 200 uL PBS. 5. Add diluted FLICA to each sample at 1:30-1:60 (e.g., spike at 1:30 by adding 10 uL to 290 uL sample). 6. Incubate approximately 1 hour. 7. Remove media and wash cells 3 times: add 1X Cellular Wash Buffer and spin cells. 8. Resuspend cell pellet in 1X Cellular Wash Buffer. 9. If desired, label with additional stains, such as Hoechst 33342, DAPI, or an antibody. 10. If desired, fix cells. 11. Analyze with a fluorescence microscope or flow cytometer. FLICA 660 is excited at 660 nm and emits at 680-690 nm. Researchers should empirically determine the suitability of the Pyroptosis/Caspase-1 Assay for an application not listed in the data sheet. Researchers commonly develop new applications and it is an integral, important part of the investigative research process. It is sometimes possible for the material contained within the vial of "Pyroptosis/Caspase-1 Assay, Assay Kit" to become dispersed throughout the inside of the vial, particularly around the seal of said vial, during shipment and storage. We always suggest centrifuging these vials to consolidate all of the liquid away from the lid and to the bottom of the vial prior to opening. Please be advised that certain products may require dry ice for shipping and that, if this is the case, an additional dry ice fee may also be required.Precautions
All products in the AAA Biotech catalog are strictly for research-use only, and are absolutely not suitable for use in any sort of medical, therapeutic, prophylactic, in-vivo, or diagnostic capacity. By purchasing a product from AAA Biotech, you are explicitly certifying that said products will be properly tested and used in line with industry standard. AAA Biotech and its authorized distribution partners reserve the right to refuse to fulfill any order if we have any indication that a purchaser may be intending to use a product outside of our accepted criteria.Disclaimer
Though we do strive to guarantee the information represented in this datasheet, AAA Biotech cannot be held responsible for any oversights or imprecisions. AAA Biotech reserves the right to adjust any aspect of this datasheet at any time and without notice. It is the responsibility of the customer to inform AAA Biotech of any product performance issues observed or experienced within 30 days of receipt of said product. To see additional details on this or any of our other policies, please see our Terms & Conditions page.Item has been added to Shopping Cart
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