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QIA108 Caspase-7 Immunoassay/Activity Kit, Fluorometric

MSDS (material safety data sheet) or SDS, CoA and CoQ, dossiers, brochures and other available documents.
Detection Methods: Fluorometric
QIA108
  
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      Overview

      Replacement Information

      Key Spec Table

      Detection Methods
      Fluorometric
      Description
      Overview

      This product has been discontinued.





      Specifically detects caspase-7 activity using the DEVD-AFC substrate. Caspase-7 substrates such as DEVD will also detect the activity of other caspases including caspase-3. This kit ensures specificity by first capturing caspase-7 using a selective antibody.
      Catalogue NumberQIA108
      Brand Family Calbiochem®
      Materials Required but Not Delivered• PBS
      References
      Product Information
      Detection methodFluorometric
      Form96 Tests
      Format96-well plate
      Kit containsCell Lsis Buffer, Coating Buffer, Anti-Caspase-7 Antibody, Blocking Buffer, Incubation Buffer, DTT, DEVD-AFC Substrate, Recombinant Caspase-7, 96-Well Plate, Plate Sealer, and a user protocol.
      Applications
      Biological Information
      Assay time6 h
      Sample TypeCultured cells
      Physicochemical Information
      Dimensions
      Materials Information
      Toxicological Information
      Safety Information according to GHS
      Safety Information
      R PhraseR: 22-38-41-52/53

      Harmful if swallowed.
      Irritating to skin.
      Risk of serious damage to eyes.
      Harmful to aquatic organisms, may cause long-term adverse effects in the aquatic environment.
      S PhraseS: 24-26-39-61

      Avoid contact with skin.
      In case of contact with eyes, rinse immediately with plenty of water and seek medical advice.
      Wear eye/face protection.
      Avoid release to the environment. Refer to special instructions/safety data sheet.
      Product Usage Statements
      Intended useThe Calbiochem® Caspase-7 Immunoassay/Activity Kit is intended for the specific, semi-quantitative detection of caspase-7 activity.
      Storage and Shipping Information
      Ship Code Dry Ice Only
      Toxicity Multiple Toxicity Values, refer to MSDS
      Storage -20°C
      Storage ConditionsUpon arrival, store the unopened kit at -20°C. Following initial thaw, store the Cell Lysis Buffer, Coating Buffer, Blocking Buffer, and Incubation Buffer at 4°C. Once opened, all components are stable for up to 6 months.
      Protect from Light Protect from light
      Avoid freeze/thaw Avoid freeze/thaw
      Do not freeze Ok to freeze
      Packaging Information
      Transport Information
      Supplemental Information
      Kit containsCell Lsis Buffer, Coating Buffer, Anti-Caspase-7 Antibody, Blocking Buffer, Incubation Buffer, DTT, DEVD-AFC Substrate, Recombinant Caspase-7, 96-Well Plate, Plate Sealer, and a user protocol.
      Specifications
      Global Trade Item Number
      Catalogue Number GTIN
      QIA108 0

      Documentation

      Brochure

      Title
      Caspases and other Apoptosis Related Tools Brochure
      Kit SourceBook - 2nd Edition EURO
      User Protocol

      Revision19-September-2008 RFH
      Form96 Tests
      Format96-well plate
      Detection methodFluorometric
      Specieshuman, mouse, rat
      StorageUpon arrival, store the unopened kit at -20°C. Following initial thaw, store the Cell Lysis Buffer, Coating Buffer, Blocking Buffer, and Incubation Buffer at 4°C. Once opened, all components are stable for up to 6 months.
      Intended useThe Calbiochem® Caspase-7 Immunoassay/Activity Kit is intended for the specific, semi-quantitative detection of caspase-7 activity.
      Principles of the assayThe Calbiochem® Caspase-7 Immunoassay/Activity Kit utilizes a caspase-7 polyclonal antibody to capture activated caspase-7 from cell lysates. A fluorogenic substrate, DEVD-AFC, is then added to the wells and is cleaved by the captured, active caspase-7. The cleavage generates free AFC that is then analyzed using a fluorescence plate reader with an excitation wavelength of 370-425 nm and an emission wavelength of 490-525 nm. The increase in fluorescence is directly proportional to the caspase-7 activity.

      Caspase-7 substrates such as DEVD will also be cleaved by other caspases, including caspase-3. However, by incorporating a specific capture step the assay only measures caspase-7 activity and other known caspases and non-specific proteases are not detected.
      Materials provided•Cell Lysis Buffer (Kit Component No. JA7541-25ML): 1 bottle, 25 ml
      •Coating Buffer (Kit Component No. JA7542-10ML): 1 bottle, 10 ml
      •Anti-Caspase-7 Antibody, 20X (Kit Component No. JA7543-500UL): 1 vial, 500 µl
      •Blocking Buffer (Kit Component No. JA7544-15ML): 1 bottle, 15 ml
      •Incubation Buffer (Kit Component No. JA7545-100ML): 1 bottle, 100 ml
      • DTT (Kit Component No. JA7546-400UL): 1 vial, 400 µl
      •DEVD-AFC Substrate (Kit Component No. JA7547-500UL): 1 vial, 500 µl
      •Recombinant Caspase-7 (Kit Component No. JA7548-10U): 1 vial, 10 U
      •96-Well Plate (Kit Component No. JA7549-EA): 1 plate, 96 wells
      •Adhesive Plate Cover (Kit Component No. JA7550-EA): 2 each
      Materials Required but not provided• PBS
      PreparationPreparation of Cell Lysates 1. Induce apoptosis as desired. Include a negative control using uninduced cells. Collect 5x106 cells per assay. 2. Wash cells with ice-cold PBS. Centrifuge at 700 x g. 3. Resuspend cells in 200 µl cold Cell Lysis Buffer. Incubate for 10 min on ice. 4. Centrifuge at 10,000 x g for 1 min. 5. Transfer the supernatant to a clean tube. Use immediately or aliquot and freeze (-20°C). The supernatant can be stored at -20°C for up to 6 months.
      Reagent preparation•1X Anti-Caspase-7 Antibody: Prepare the desired amount of 1X Anti-Caspase-7 Antibody by making a 1:20 dilution of the Anti-Caspase-7 Antibody, 20X in Coating Buffer. •Recombinant Caspase-7: Reconstitute the Recombinant Caspase-7 in 10 µl PBS. Dilute 1:100 in Cell Lysis Buffer as needed for the assay.
      Detailed protocol1. Add 100 µl 1X Anti-Caspase-7 Antibody to each well. Cover the plate tightly with an Adhesive Plate Cover and incubate at 37°C for 1 h or at 4°C overnight.
      2. Remove the antibody solution from the wells.
      3. Add 150 µl Blocking Buffer to each well. Cover the plate tightly with an Adhesive Plate Cover and incubate at room temperature for 30 min.
      4. Remove the Blocking Buffer and wash the wells 3 times with 150 µl Incubation Buffer.
      5. Add 100 µl cell lysate or 1 unit Recombinant Caspase-7 (positive control) to Anti-Caspase-7 Antibody-coated wells. Cover the plate tightly with an Adhesive Plate Cover and incubate at 37°C for 1 h.
      6. Remove the solutions from the wells and wash 3 times with 150 µl Incubation Buffer.
      7. Add 94 µl Incubation Buffer, 5 µl DEVD-AFC, and 1 µl DTT to each well. Cover the plate tightly with an Adhesive Plate Cover and incubate for 2-4 h at 37°C. Overnight incubation at 37°C may increase sensitivity.
      8. Read the samples using a fluorescence plate reader set at an excitation wavelength of 370-425 nm and an emission wavelength of 490-525 nm. Determine the Caspase-7 activity by comparing results with the activity level of the uninduced control.
      Registered TrademarksCalbiochem® is a registered trademark of EMD Biosciences, Inc.
      Interactive Pathways™ is a trademark of EMD Biosciences, Inc.