FLICA™ Caspase 1 Detection Kit

Quantitate caspase 1 (ICE) in vitro with FAM-YVAD-FMK

ICT's FLICA caspase 1 detection assay is a simple yet accurate method to measure caspase 1 (ICE) activity in whole cells. Simply add the FLICA caspase 1 reagent to the media and incubate. Cells containing active caspase 1 will fluoresce green. The kit contains:

ICT's FLICA caspase 1 detection probe, FAM-YVAD-FMK
10x apoptosis wash buffer
Hoechst stain (a blue DNA stain)
Propidium iodide (a red live/dead stain)
Fixative (for use after labeling with FLICA)

Trial size kit (~25 tests), cat. no. 97, $169

Regular size kit (~100 tests), cat. no. 98, $473

Caspase 1 activity may be detected in individual cells or suspensions of up to 1x10^6 cells/mL. The noncytotoxic FLICA probe is cell permeant: no lysis or permeabilization steps are necessary. Just culture cells, add FLICA to the media, incubate, and wash.

Active caspase 1 enzymes will bind to FLICA and retain it within the cells, capturing the green fluorescent signal.

FLICA constantly fluoresces, therefore any unbound reagent must be washed out of the cell to remove any background noise.

The resulting positive fluorescent signal can be detected with a fluorescence microscope, a fluorescence plate reader, or a flow cytometer.

The green FLICA probe, FAM, excites at 490nm and emits at 520nm.

The FLICA assay also works with adherent cells and very thin tissue sections. FLICA will not work in previously fixed cells or paraffin cells, as those treatments inactivate the caspase 1 enzyme. Please contact us for technical assistance.

ICT's FLICA caspase 1 inhibitor probe is comprised of 3 segments:

a green fluorescent label, FAM (carboxyfluorescein)
an inhibitor peptide sequence targeted by active caspase 1 (YVAD)
and a fluoromethylketone group (FMK), which forms a covalent bond with the active enzyme.

FLICA kits have been used successfully with a multitude of cell lines from a variety of species (human, mouse, rat, etc), including, but not limited to, cancer cells, neurons, primary cells, epithelial cells, endothelial cells, and macrophages. For in vitro apoptosis detection via active poly caspases, see our FLICA Poly Caspases Detection Kits.

Fluorescent Probes Application Guide 2010 Price List Order Form

Cells were treated with an inducing agent, or DMSO, a negative control, for 4 hours, then labeled with ICT’s green caspase 1 inhibitor probe, FAM-YVAD-FMK, for 1 hour and examined under a fluorescence microscope. The images to the lower left reveal 2 experimental cells, both of which fluoresce green, therefore they have active caspases. The brightfield image to the lower right reveals many control cells in the field of view, however the corresponding fluorescence image is dark, therefore, none of these cells have active caspases (Dr. Brian W. Lee, ICT).

Caspase activity

	Brochure		Papers
	Manual		References
	MSDS

Sample protocol:

Culture your cells up to 1 x 10^6 cells/mL.
Induce apoptosis following your protocol, and create positive and negative controls.
Reconstitute the reagent with 50mcL DMSO to form the stock concentrate (which can be frozen for future use).
Dilute the stock concentrate with 200mcL 1X PBS to form the working solution.
Add ~10mcL of the working solution directly to a 300-500mcL aliquot of your cell culture for labeling.
Incubate 1-4 hours.
Wash and spin cells twice, or let incubate for 1 hour with fresh media or 1x apoptosis wash buffer.
If desired, label cells with Hoechst stain.
If desired, label cells with Propidium Iodide or 7AAD.
If desired, fix cells.
Analyze data using a fluorescence microscope, plate reader, or flow cytometer.