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  • 产品名称：Caspase-10SubstrateAEVD-pNA

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  • 产品厂商：Biovision
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简单介绍：
Caspase-10SubstrateAEVD-pNA
详情介绍：
Sequence Ac-Ala-Glu-Val-Asp-pNA
Characteristics Ready-to-use colorimetric substrate for caspases that recognize the amino acid sequence AEVD. Caspase activity can be quantified by spectrophotometric detection of free pNA (l = 400 nm) after cleaved from the peptide substrate AEVD-pNA, using a spectrophotometer or multi-well plate reader. The ready-to-use caspase substrate provides an economical alternative for researchers who perform large volume of caspase assays.
Purity > 99 % by HPLC
Chemical Name Ac-AEVD-pNA, Caspase-10 Substrate, Colorimetric
Formula C₂₅H₃₄N₆O₁₁
Permeability Not-permeable
Molecular Weight 594.7 g/mol
Comment

Note: Background reading from cell lysates and buffers should be subtracted from the readings of both induced and the uninduced samples before calculating fold increase in caspase activity.
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.
3. Resuspend cells in 50 µL of chilled Cell Lysis Buffer and incubate cells on ice for 10 minutes.
4. Centrifuge for 1 min in a microcentrifuge (10,000 x g).
5. Transfer supernatant (cytosolic extract) to a fresh tube and put on ice.
6. Assay protein concentration.
7. Dilute 100-200 µg protein to 50 µL Cell Lysis Buffer for each assay.
8. Add 50 µL of 2X Reaction Buffer containing 10 mM DTT to each sample.
9. Add 5 µL of the 4 mM of AEVD-pNA (200 µ M final conc.) and incubate at 37 °C for 1-2 hour.
10. Read samples at 400- or 405-nm in a microtiter plate reader or spectrophotometer using a 100- µL micro quartz cuvet (Sigma), or dilute sample to 1 mL with Dilution Buffer and using regular cuvet (note: Dilution of the samples proportionally decreases the reading). Fold-increase in caspase activity can be determined by comparing these results with the level of the uninduced control. Note: Background reading from cell lysates and buffers should be subtracted from the readings of both induced and the uninduced samples before calculating fold increase in caspase activity.
Restrictions For Research Use only
Format Liquid
Handling Advice Protect from light and moisture
Storage -20 °C
Expiry Date 12 months
Product cited in: Bian, Sun, Silver, Ho, Marchionni, Caggiano, Stone, Amende, Hampton, Morgan, Yan: "Neuregulin-1 attenuated doxorubicin-induced decrease in cardiac troponins." in: American journal of physiology. Heart and circulatory physiology, Vol. 297, Issue 6, pp. H1974-83, 2009 (PubMed).

Doostzadeh-Cizeron, Yin, Goodrich: "Apoptosis induced by the nuclear death domain protein p84N5 is associated with caspase-6 and NF-kappa B activation." in: The Journal of biological chemistry, Vol. 275, Issue 33, pp. 25336-41, 2000 (PubMed).