Caspase-2 Substrate VDVAD-AFC

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Enzyme Activity Assay (EAA), Functional Studies (Func)
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Sequence Ac-Val-Asp-Val-Ala-Asp-AFC
Characteristics Ready-to-use fluorometric substrate for caspases that recognize the amino acid sequence VDVAD. Caspase activity can be quantified by fluorescent detection of free AFC after cleaved from the peptide substrate VDVAD-AFC at Ex/Em = 400/505 nm using a fluorometer or a fluorescence plate reader. Alternatively, a shift in fluorescence from blue to green upon cleavage can be visualized using a hand-held long-UV lamp. The ready-to-use caspase substrate provides an economic alternative for researchers who perform large amount of caspase assays. Cell Lysis Buffer, and other reagents used for caspase activity assays are also available separately.
Purity > 98 % by HPLC
Chemical Name Ac-VDVAD-AFC, Caspase-2 Substrate, Fluorogenic, VDVAD-AFC, caspase2 substrate, caspase 2 substrate
Formula C₃₃H₄₁F₃N₆O₁₂
Permeability Not-permeable
Molecular Weight 770.7 g/mol
Protocol 1. Induce apoptosis in cells by desired method. Concurrently incubate a control culture without induction.
2. Count cells and pellet 1-5 x 10^6 cells or use 50-200 µg cell lysates if protein concentration has been measured.
3. Resuspend cells in 50 µL of chilled Cell Lysis Buffer containing 10 mM DTT to each sample.
6. Add 5 µL of the 1 mM VDVAD-AFC (50 µ M final conc.) into each tube individually and incubate at 37 °C for 1-2 hour.
7. Read samples in a fluorometer equipped with a 400-nm excitation filter and 505-nm emission filter. For a plate-reading set-up, transfer the samples to a 96-well plate. You may perform the entire assay directly in a 96-well plate. Fold-increase in VDVAD-dependent caspase activity can be determined by comparing these results with the level of the uninduced control.
Restrictions For Research Use only
Format Liquid
Handling Advice Protect from light and moisture
Storage -20 °C
Expiry Date 12 months
Product cited in: Basu, Adkins, Basu: "Down-regulation of caspase-2 by rottlerin via protein kinase C-delta-independent pathway." in: Cancer research, Vol. 68, Issue 8, pp. 2795-802, 2008 (PubMed).

Ray, Akbiyik, Bernstein, Phipps: "CD40 engagement prevents peroxisome proliferator-activated receptor gamma agonist-induced apoptosis of B lymphocytes and B lymphoma cells by an NF-kappaB-dependent mechanism." in: Journal of immunology (Baltimore, Md. : 1950), Vol. 174, Issue 7, pp. 4060-9, 2005 (PubMed).

Bai, Goodrich: "Different DNA lesions trigger distinct cell death responses in HCT116 colon carcinoma cells." in: Molecular cancer therapeutics, Vol. 3, Issue 5, pp. 613-9, 2004 (PubMed).

Li, Chen, Sinogeeva, Gorospe, Wersto, Chrest, Barnes, Liu: "Arsenic trioxide promotes histone H3 phosphoacetylation at the chromatin of CASPASE-10 in acute promyelocytic leukemia cells." in: The Journal of biological chemistry, Vol. 277, Issue 51, pp. 49504-10, 2002 (PubMed).