Caspase-6 Substrate VEID-AFC

Details for Product No. ABIN412382
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Application
Functional Studies (Func), Enzyme Activity Assay (EAA)
Pubmed 3 references available
Quantity 1000 tests
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Catalog No. ABIN412382
599.50 $
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Sequence Ac-Val-Glu-Ile-Asp-AFC
Purity > 95 % by HPLC
Molecular Weight 727.7 Da
Comment

Ready-to-use fluorometric substrate for caspase-6 and related caspases that recognize the amino acid sequence VEID. Caspase-6 and related caspase activity can be quantified by fluorescent detection of free AFC after cleavage from the peptide substrate VEID-AFC at Ex/Em = 400/505, using a fluorometer or multi-well 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-6 assays. Cell Lysis Buffer for caspase assays are also available separately.

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 VEID-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 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: Coletti, Yang, Marazzi et al.: "TNFalpha inhibits skeletal myogenesis through a PW1-dependent pathway by recruitment of caspase pathways." in: The EMBO journal, Vol. 21, Issue 4, pp. 631-42, 2002 (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).

Deane, Trifilo, Yballe et al.: "Enhanced T cell proliferation in mice lacking the p85beta subunit of phosphoinositide 3-kinase." in: Journal of immunology (Baltimore, Md. : 1950), Vol. 172, Issue 11, pp. 6615-25, 2004 (PubMed).

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