Active caspase-1 was detected in living ECs with use of the Fluorescent Labeled Inhibitor of Caspases (FLICA) caspase-1 assay kit (ImmunoChemistry Technologies). FLICA-FAM-YVAD-FMK is a cell-permeable, non-toxic fluorochrome inhibitor of caspase-1 that interacts with the enzymatic-reactive center of activated caspase-1 via the YVAD recognition sequence, thus forming a covalent thioether adduct with the enzyme through the FMK moiety. The resulting green fluorescence is a direct measure of caspase-1 activity, which was analyzed by FACS with 488-nm excitation and 530-nm emission. NO was detected as the accumulated nitrite/nitrate, the stable breakdown product of NO, in cell culture media by use of a nitrate/nitrite fluorometric assay (Cayman Chemicals).
Flica caspase 1 assay kit
Manufactured by ImmunoChemistry Technologies
Sourced in United States
The FLICA (Fluorochrome Inhibitors of Caspases) caspase-1 assay kit is a laboratory tool used to detect and measure the activity of caspase-1, an enzyme involved in the inflammatory response. The kit utilizes a fluorescent-labeled inhibitor that binds to active caspase-1, allowing for quantification of the enzyme's presence and activity in biological samples.
Automatically generated - may contain errors
3 protocols using flica caspase 1 assay kit
1
Caspase-1 and Nitric Oxide Detection
2
Caspase Activation in J774 Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
J774 cells (2×104 cells/well) were seeded in CC2 Chamber Slide system 8-wells (Thermo Fisher Scientific) overnight, and then treated with NETs (1.7×105 cell equivalents) for 24 h in the absence or presence of 10 ng/ml LPS at 37°C in 5% CO2. Thereafter, the activation of caspase-1 and caspase-8 was assayed using the FLICA™ caspase-1 assay kit (ImmunoChemistry Technologies, LLC, Bloomington, MN, USA) and FLICA™ caspase-8 assay kit (ImmunoChemistry Technologies, LLC), respectively. Briefly, after treatment with LPS and NETs, the cells were washed with fresh RPMI-1640 medium, and then the cells were labeled with FAM-YVAD-FMK or FAM-LETD-FMK (fluorescent-labeled caspase-1 or caspase-8 inhibitor that binds with activated caspase-1 or caspase-8, respectively) by incubation for 1 h at 37°C. After washing, the cells were labeled with Hoechst 33342 (0.8 µg/ml) for 10 min. Finally, the slide was mounted with a cover glass (Paul Marienfeld GmbH & Co. KG, Lauda-Königshofen, Germany) using aqueous medium Vectashield, and the cells were photo-graphed using a fluorescence microscope system Axioplan 2.
3
LL-37 Modulates Caspase-1 Activation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
J774 cells (1×106 cells/well) in 6 well-culture plates were primed with 10 ng/ml LPS for 4 h in the absence or presence of LL-37 (0.01, 0.1 or 1 µg/ml), and then treated with 3 mM ATP for 90 min. Thereafter, the caspase-1 activation was assayed by using a FLICA™ Caspase-1 Assay Kit (Immunochemistry Technologies, Bloomington, MN). Briefly, after the treatment, cells were detached using cell scrapers, washed and suspended in 300 µl DMEM. Then, the cells were labeled with FAM-YVAD-fmk (fluorescent labeled caspase-1 inhibitor that bind with activated caspase-1) by incubation for 1 h at 37°C, washed and finally assayed by flow cytometry, according to the manufacturer's manual using FACSCalibur (BD Biosciences, Rutherford, NJ, USA). In separate experiments, J774 cells were directly treated with ATP in the presence or absence of LL-37 or P2X7 antagonist (KN-62 or KN-93), and the caspase-1 activation was evaluated, as described above.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!