货号：APT172

品牌：Merck Millipore

规格：EA

目录价：￥6891.00

市场价格：￥5857.35

会员价格：￥5512.80

金山科研平台，产品价格货期咨询微信：jinshanbio Description: Caspase 9 Fluorometric Activity Assay Kit, LEHD View All» Trade Name: Chemicon (Millipore) View All» Qty/Pk: 100 assays View All» Product Overview: Activation of ICE-family proteases/caspases initiates apoptosis in mammalian cells. Chemicon's MCH6/Caspase-9 Fluorometric Protease Assay Kit provides a simple and convenient means for assaying the activity of caspases that recognize the sequence LEHD. The assay is based on detection of cleavage of substrate LEHD-AFC (7-amino-4-trifluoromethyl coumarin). LEHD-AFC emits blue light (lmax=400 nm); upon cleavage of the substrate by MCH-6 or related caspases, free AFC emits a yellow-green fluorescence (lmax=505 nm), which can be quantified using a fluorometer or a fluorescence microtiter plate reader. Comparison of the fluorescence of AFC from an apoptotic sample with an uninduced control allows determination of the fold increase in Caspase-9 activity. For research use only; Not for use in diagnostic procedures View All» Key Applications: Activity Assay View All» Application Notes: Preparation of Reagents and Assay Instructions General Considerations: • Aliquot enough 2X Reaction Buffer for the number of assays to be performed. Add DTT to the 2X Reaction Buffer immediately before use (10 mM final concentration: add 10 μL of 1.0 M DTT stock per 1 mL of 2X Reaction Buffer). • After thawing, store the Cell Lysis Buffer and 2X Reaction Buffer at 4oC. • Protect LEHD-AFC from light. Assay Procedure: 1. Induce apoptosis in cells by desired method. Concurrently incubate a control culture without induction. 2. Count cells and pellet 1-5 X 106 cells (1500 rpm for 10 minutes) or use 50- 200 μg cell lysates if protein concentration has been measured. 3. Resuspend cells in 50 μL of chilled Cell Lysis Buffer. 4. Incubate cells on ice for 10 minutes. 5. Add 50 μL of 2X Reaction Buffer (containing 10 mM DTT) to each sample. 6. Add 5 μL of the 1 mM LEHD-AFC substrate (50 μM final concentration) and incubate at 37°C for 1-2 hours. 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 also perform the entire assay directly in a 96-well plate. Fold-increase in Caspase-9 activity can be determined by comparing these results with the level of the uninduced control. View All» Species Reactivity: Mammals View All» Usage Statement: Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals. View All» Entrez Gene Summary: This gene encodes a member of the cysteine-aspartic acid protease (caspase) family. Sequential activation of caspases plays a central role in the execution-phase of cell apoptosis. Caspases exist as inactive proenzymes which undergo proteolytic processing at conserved aspartic residues to produce two subunits, large and small, that dimerize to form the active enzyme. This protein is processed by caspase APAF1; this step is thought to be one of the earliest in the caspase activation cascade. Alternative splicing results in two transcript variants which encode different isoforms. View All» UniProt Summary: FUNCTION: SwissProt: P55211 # Isoform 2 lacks activity is an dominant-negative inhibitor of caspase-9.SIZE: 416 amino acids; 46281 Da SUBUNIT: Heterotetramer that consists of two anti-parallel arranged heterodimers, each one formed by a 35 kDa (p35) and a 10 kDa (p10) subunit. Caspase-9 and APAF1 bind to each other via their respective NH2-terminal CED-3 homologous domains in the presence of cytochrome C and ATP. Interacts with the inhibitors BIRC2, BIRC4, BIRC5 and BIRC7.TISSUE SPECIFICITY: Ubiquitous, with highest expression in the heart, moderate expression in liver, skeletal muscle, and pancreas. Low levels in all other tissues.PTM: Cleavages at Asp-315 by granzyme B and at Asp-330 by caspase- 3 generate the two active subunits. Caspase-8 and -10 can also be involved in these processing events.SIMILARITY: SwissProt: P55211 ## Belongs to the peptidase C14 family. & Contains 1 CARD domain. View All» Components:

• Cell Lysis Buffer - 100 mL

• 2X Reaction Buffer - 4 x 2 mL

• DTT (1 M): 400 μL

• LEHD-AFC Substrate: 500 μL
  View All» Brand Family: Chemicon View All» UniProt Number: P55211 View All» Storage Conditions: We recommend storing reagents at -20°C upon arrival. (Store Cell Lysis Buffer and 2X Reaction Buffer at 4°C after opening) upon arrival. The performance of this product is guaranteed for one year under proper storage conditions. View All» Detection Methods: Fluorescent View All» Gene Symbol:
  • CASP9

  • CASP-9

  • ICE-LAP6

  • MCH6

  • APAF3

  • CASPASE-9c

  • OTTHUMP00000044594

  • APAF-3

  • EC 3.4.22.62 [Contains: Caspase-9 subunit p35

  • Caspase-9 subunit p10].
    View All» Product Name: Caspase 9 Fluorometric Activity Assay Kit, LEHD View All» Entrez Gene Number:
    • NM_001229.2

    • NM_032996.1
      View All» Alternate Names: Mch-6; ICE-LAP6; Apaf-3 View All» Materials Required but Not Delivered: · Microcentrifuge and 1.5 mL Microcentrifuge tubes · 37°C water bath or incubator · Fluorometer equipped with a 400-nm excitation filter and 505-nm emission filter. · 96 well plate View All»
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