Z-IETD-AFC *CAS 219138-02-0*
Z-IETD-AFC is a fluorogenic caspase-8/granzyme B substrate containing the acetyl (Ac) moiety. This substrate is hydrolyzed by caspase 8 to generate highly fluorescent 7-amido-4-trifluoromethylcoumarin (AFC).
Example protocol
AT A GLANCE
Important notes
It is important to store at <-15 °C and should be stored in cool, dark place.
It can be used within 12 months from the date of receipt.
SAMPLE EXPERIMENTAL PROTOCOL
Following protocol only provides a guideline, and should be modified according to your specific needs.
General Solution Caspase Assays Using AMC, AFC, pNA, R110 and ProRed Substrates
- Prepare a 10 mM stock solution in DMSO.
- Prepare a 2X caspase substrate (50 µM) assay solution as the following: 50 µL substrate stock solution, 100 µL DTT (1M), 400 µL EDTA (100 mM), 10 mL Tris Buffer (20 mM), pH =7.4.
- Mix equal volume of the caspase standards or samples with 2X caspase substrate assay solution, and incubate the solutions at room temperature for at least 1 hour.
- Monitor the fluorescence using a fluorescence microplate reader, or absorbance using an absorbance microplate reader.
Cell Caspase Assays Using Cell-Permeable FMK Caspase Probes
- Prepare a 2-5 mM stock solution in DMSO.
- Treat cells as desired.
- Prepare a 2X permeable caspase substrate (20 µM) assay solution by diluting the DMSO stock solution (from Step 2.1) in Hanks with 20 mM Hepes buffer (HHBS).
- Mix equal volume of the treated cells with 2X caspase substrate assay solution (from Step 2.3), and incubate the cells in a 37°C, 5% CO2 incubator for at least1 hour.
- Wash the cells with HHBS for at least once.
- Monitor the fluorescence intensity by a flow cytometer, a fluorescence microscope or a fluorescence microplate reader.
Cell Caspase Assays Using Cell-Permeable FMK Caspase Probes (For #13470-13476 only)
- Prepare a 250X stock solution by adding 50 µL DMSO into the vial.
- Treat cells as desired.
- Add 250 X DMSO stock solution into the cell solution at a 1:250 ratio (such as 2 µL to 500 µL cells), and incubate the cells in a 37°C, 5% CO2 incubator for 1 hour.
- Wash the cells with HHBS for at least once.
- Monitor the fluorescence intensity by flow cytometer, fluorescence microscopy or fluorescent microplate reader.
Calculators
Common stock solution preparation
Table 1. Volume of DMSO needed to reconstitute specific mass of Z-IETD-AFC *CAS 219138-02-0* to given concentration. Note that volume is only for preparing stock solution. Refer to sample experimental protocol for appropriate experimental/physiological buffers.
0.1 mg | 0.5 mg | 1 mg | 5 mg | 10 mg | |
1 mM | 121.692 µL | 608.458 µL | 1.217 mL | 6.085 mL | 12.169 mL |
5 mM | 24.338 µL | 121.692 µL | 243.383 µL | 1.217 mL | 2.434 mL |
10 mM | 12.169 µL | 60.846 µL | 121.692 µL | 608.458 µL | 1.217 mL |
Molarity calculator
Enter any two values (mass, volume, concentration) to calculate the third.
Mass (Calculate) | Molecular weight | Volume (Calculate) | Concentration (Calculate) | Moles | ||||
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Spectrum
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Product family
Name | Excitation (nm) | Emission (nm) | Extinction coefficient (cm -1 M -1) |
Ac-IETD-AFC *CAS 211990-57-7* | 376 | 482 | 170001 |
Z-DEVD-AFC | 376 | 482 | 170001 |
Z-IETD-aminoluciferin | 362 | 499 | 18000 |
Citations
View all 2 citations: Citation Explorer
ONC206, an imipridone derivative, induces cell death through activation of the integrated stress response in serous endometrial cancer in vitro
Authors: Zhang, Yingao and Huang, Yu and Yin, Yajie and Fan, Yali and Sun, Wenchuan and Zhao, Xiaoling and Tucker, Katherine and Staley, Allison and Paraghamian, Sarah and Hawkins, Gabrielle and others,
Journal: Frontiers in Oncology (2020): 577141
Authors: Zhang, Yingao and Huang, Yu and Yin, Yajie and Fan, Yali and Sun, Wenchuan and Zhao, Xiaoling and Tucker, Katherine and Staley, Allison and Paraghamian, Sarah and Hawkins, Gabrielle and others,
Journal: Frontiers in Oncology (2020): 577141
pH-Assisted surface functionalization of selenium nanoparticles with curcumin to achieve enhanced cancer chemopreventive activity
Authors: Yu, Shaoxuan and Wang, Yanru and Zhang, Wentao and Zhang, Yuhuan and Zhu, Wenxin and Liu, Yingnan and Zhang, Daohong and Wang, Jianlong
Journal: RSC Advances (2016): 72213--72223
Authors: Yu, Shaoxuan and Wang, Yanru and Zhang, Wentao and Zhang, Yuhuan and Zhu, Wenxin and Liu, Yingnan and Zhang, Daohong and Wang, Jianlong
Journal: RSC Advances (2016): 72213--72223
References
View all 67 references: Citation Explorer
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Measurement of two caspase activities simultaneously in living cells by a novel dual FRET fluorescent indicator probe
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Authors: Wu X, Simone J, Hewgill D, Siegel R, Lipsky PE, He L.
Journal: Cytometry A (2006): 477
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Photoreceptor cell apoptosis induced by the 2-nitroimidazole radiosensitizer, CI-1010, is mediated by p53-linked activation of caspase-3
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Diallyl Trisulfide Induces Apoptosis of Human Gastric Cancer Cell Line MGC803 Through Caspase-3 Pathway.
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Authors: Xiao XL, Peng J, Su Q, Xiang SL, Tang GH, Huang YS, Zhou XT.
Journal: Ai Zheng (2006): 1247
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