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Z-VAD-FMK

Inhibition of caspase activities in Jurkat cells with pan caspase inhibitor Z-VAD-FMK (Cat.# 13300). Jurkat cells were seeded on the same day at 200,000 cells/90 µL/well in a Costar black wall/clear bottom 96-well plate. The cells were treated with or without 1 µM of staurosporine for 5 hours. The Z-VAD-FMK (50 uM) was added and co-incubated with staurosporine. The caspase-8 activity was detected with Cell Meter™ Caspase 8 Activity Apoptosis Assay Kit (Cat.# 22816). The fluorescence intensity was measured at Ex/Em = 540/620 nm (Cutoff = 610 nm) with Flexstation 3 fluorescence microplate reader (Molecular Devices).
Inhibition of caspase activities in Jurkat cells with pan caspase inhibitor Z-VAD-FMK (Cat.# 13300). Jurkat cells were seeded on the same day at 200,000 cells/90 µL/well in a Costar black wall/clear bottom 96-well plate. The cells were treated with or without 1 µM of staurosporine for 5 hours. The Z-VAD-FMK (50 uM) was added and co-incubated with staurosporine. The caspase-8 activity was detected with Cell Meter™ Caspase 8 Activity Apoptosis Assay Kit (Cat.# 22816). The fluorescence intensity was measured at Ex/Em = 540/620 nm (Cutoff = 610 nm) with Flexstation 3 fluorescence microplate reader (Molecular Devices).
Chemical structure for Z-VAD-FMK.
Ordering information
Price ()
Catalog Number13300
Unit Size
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Additional ordering information
Telephone1-408-733-1055
Fax1-408-733-1304
Emailsales@aatbio.com
InternationalSee distributors
ShippingStandard overnight for United States, inquire for international
Physical properties
Molecular weight467.49
SolventDMSO
Storage, safety and handling
H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22
StorageFreeze (< -15 °C); Minimize light exposure
UNSPSC12171501

OverviewpdfSDSpdfProtocol


Molecular weight
467.49
Z-VAD-FMK (carbobenzoxy-valyl-alanyl-aspartyl-[O-methyl]- fluoromethylketone) is a cell-permeable, irreversible pan caspase inhibitor that binds to the catalytic site of caspase proteases and can inhibit induction of apoptosis. For inhibition of apoptosis, Z-VAD-FMK should be added at the same time that apoptosis is induced. The peptide is O-methylated in the P1 position on aspartic acid, providing enhanced stability and increased cell permeability. Fluoromethyl ketone (FMK)-derivatized peptides act as effective irreversible inhibitors with no added cytotoxic effects.

Example protocol


PREPARATION OF STOCK SOLUTIONS

Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles.

Z-VAD-FMK stock solution
Add appropriate amount of DMSO into Z-VAD-FMK to make 2-5 mM Z-VAD-FMK stock solution.
Note     Store the unused Z-VAD-FMK stock solution at -20 °C in single use aliquots.

SAMPLE EXPERIMENTAL PROTOCOL

The following protocol can be used as a guideline and should be optimized according to your needs.
  1. Treat your samples as desired to induce apoptosis.
  2. Add Z-VAD-FMK along with the treatment of your choice.
    Note     We recommend using 10-100 µM of final concentration ofZ-VAD-FMK to inhibit caspases such as 50 µM for caspase-8.
    Note     Optimal time and concentration for incubation needs to be determined experimentally.
  3. Stain samples as you desired.
  4. Monitor the fluorescence intensity with a proper instrument. 

Calculators


Common stock solution preparation

Table 1. Volume of DMSO needed to reconstitute specific mass of Z-VAD-FMK to given concentration. Note that volume is only for preparing stock solution. Refer to sample experimental protocol for appropriate experimental/physiological buffers.

0.1 mg0.5 mg1 mg5 mg10 mg
1 mM213.908 µL1.07 mL2.139 mL10.695 mL21.391 mL
5 mM42.782 µL213.908 µL427.817 µL2.139 mL4.278 mL
10 mM21.391 µL106.954 µL213.908 µL1.07 mL2.139 mL

Molarity calculator

Enter any two values (mass, volume, concentration) to calculate the third.

Mass (Calculate)Molecular weightVolume (Calculate)Concentration (Calculate)Moles
/=x=

Product family


NameExcitation (nm)Emission (nm)
FAM-VAD-FMK493517
TF4-VAD-FMK578602
SRB-VAD-FMK [Sulforhodamine B-VAD-FMK]559577
Z-DEVD-FMK--
Z-IETD-FMK--
Z-LETD-FMK--

References


View all 8 references: Citation Explorer
Identification of active caspases using affinity-based probes.
Authors: McStay, Gavin P and Green, Douglas R
Journal: Cold Spring Harbor protocols (2014): 856-60
Induction of apoptosis in human renal cell carcinoma cells by vitamin E succinate in caspase-independent manner.
Authors: Wu, Xiu-Xian and Kakehi, Yoshiyuki and Jin, Xing-Hua and Inui, Masashi and Sugimoto, Mikio
Journal: Urology (2009): 193-9
Metformin induces apoptosis of pancreatic cancer cells.
Authors: Wang, Luo-Wei and Li, Zhao-Shen and Zou, Duo-Wu and Jin, Zhen-Dong and Gao, Jun and Xu, Guo-Ming
Journal: World journal of gastroenterology (2008): 7192-8
Induction of apoptosis in renal tubular cells by histone deacetylase inhibitors, a family of anticancer agents.
Authors: Dong, Guie and Wang, Lysa and Wang, Cong-Yi and Yang, Tianxin and Kumar, M Vijay and Dong, Zheng
Journal: The Journal of pharmacology and experimental therapeutics (2008): 978-84
Apoptosis of gastric cancer cell line MKN45 by photodynamic treatment with photofrin.
Authors: Takahira, Kenichiro and Sano, Munetaka and Arai, Hajime and Hanai, Hiroyuki
Journal: Lasers in medical science (2004): 89-94
Non-caspase-mediated apoptosis contributes to the potent cytotoxicity of the enediyne antibiotic lidamycin toward human tumor cells.
Authors: Wang, Zhen and He, Qiyang and Liang, Yunyan and Wang, Daishu and Li, Yi Yang and Li, Diandong
Journal: Biochemical pharmacology (2003): 1767-75
Androgen induced cell death in SHSY5Y neuroblastoma cells expressing wild-type and spinal bulbar muscular atrophy mutant androgen receptors.
Authors: Grierson, A J and Shaw, C E and Miller, C C
Journal: Biochimica et biophysica acta (2001): 13-20
Critical role for caspases 3 and 8 in neutrophil but not eosinophil apoptosis.
Authors: Daigle, I and Simon, H U
Journal: International archives of allergy and immunology (2001): 147-56