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QIA114 Caspase-8 Detection Kit (Red-IETD-FMK)

Caspase-8 Detection Kit (Red-IETD-FMK) MSDS (material safety data sheet) or SDS, CoA and CoQ, dossiers, brochures and other available documents.
QIA114
  
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      Overview

      Replacement Information

      Key Spec Table

      Detection Methods
      Fluorescence
      Description
      Overview

      This product has been discontinued.





      A fast, convenient, and sensitive method to measure caspase-8 in living cells. The fluorescent marker, Red-IETD-FMK, is a labeled, cell-permeable, non-toxic inhibitor that binds irreversibly to activated caspase-8 in living cells. Caspase-8 can be detected by the measurement of fluorescence intensity in a fluorescent microplate reader, flow cytometer, or by fluorescence microscopy.
      Catalogue NumberQIA114
      Brand Family Calbiochem®
      References
      Product Information
      Detection methodFluorescence
      Form100 Tests
      FormatFlow cytometry or fluorescence microscopy
      Kit containsLabeled Caspase-8 Inhibitor (Red-IETD-FMK), Wash Buffer, Unlabeled Caspase Inhibitor (Z-VAD-FMK), and a user protocol.
      Applications
      Biological Information
      Assay time1.5 h
      Sample TypeIntact cells
      Physicochemical Information
      Emission max.
      Excitation max.
      Dimensions
      Materials Information
      Toxicological Information
      Safety Information according to GHS
      Safety Information
      Product Usage Statements
      Storage and Shipping Information
      Ship Code Dry Ice Only
      Toxicity Multiple Toxicity Values, refer to MSDS
      Storage -20°C
      Storage ConditionsUpon arrival store the entire contents of the kit at -20°C.
      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 containsLabeled Caspase-8 Inhibitor (Red-IETD-FMK), Wash Buffer, Unlabeled Caspase Inhibitor (Z-VAD-FMK), and a user protocol.
      Specifications
      Global Trade Item Number
      Catalogue Number GTIN
      QIA114 0

      Documentation

      Caspase-8 Detection Kit (Red-IETD-FMK) Certificates of Analysis

      TitleLot Number
      QIA114
      User Protocol

      Revision10-March-2010 RFH
      Form100 Tests
      FormatFlow cytometry or fluorescence microscopy
      Detection methodFluorescence
      Speciesa broad range of species
      StorageUpon arrival store the entire contents of the kit at -20°C.
      Principles of the assayThe activation of caspases plays a central role in apoptosis. This Calbiochem® brand Caspase-8 Detection Kit provides a convenient and sensitive means for detecting activated caspases in living cells. The fluorescent marker, Red-IETD-FMK, is a labeled, cell-permeable, non-toxic inhibitor that binds irreversibly to activated caspase-8 in living cells. Caspase-8 can then be detected by the measurement of fluorescence intensity (excitation max.: ~540 nm; emission max.: ~570 nm) in a fluorescent microplate reader, by flow cytometry, or by fluorescence microscopy.
      Materials provided• Red-IETD-FMK (Kit Component No. JA7623) 1 vial, 100 µl
      • Wash Buffer (Kit Component No. JA7624) 2 bottles, 100 ml
      • Z-VAD-FMK (Kit Component No. JA7625) 1 vial, 10 µl
      Detailed protocolStaining

      1. Induce apoptosis in cells (1 x 106/ml) as desired. Concurrently incubate a control culture without induction. An additional negative control can be prepared by adding the caspase inhibitor Z-VAD-FMK at 1 µl/ml to an induced culture to inhibit caspase activation.
      2. Aliquot 300 µl each of the induced and control cultures into eppendorf tubes.
      3. Add 1 µl of Red-IETD-FMK to each tube and incubate for 0.5-1 h in a 37°C incubator with 5% CO2.
      4. Centrifuge cells at 3,000 rpm for 5 min and remove supernatant.
      5. Resuspend cells in 0.5 ml of Wash Buffer, and centrifuge again.
      6. Repeat Step 5.

      Proceed depending on method of analysis.

      Quantification by Flow Cytometry

      1. For flow cytometric analysis, resuspend cells in 300 µl of Wash Buffer and place on ice.
      2. Analyze samples by flow cytometry using the FL-2 channel.

      Detection by Fluorescence Microscopy

      1. Resuspend cells in 100 µl of Wash Buffer.
      2. Add one drop of the cell suspension to a microslide and cover with a coverslip.
      3. Observe cells under a fluorescence microscope using a rhodamine filter. Caspase-8 positive cells appear to have bright red signals, whereas caspase-8 negative control cells show a much weaker signal.

      Analysis by Fluorescence Plate Reader

      1. Resuspend cells in 100 µl of Wash Buffer.
      2. Transfer the cell suspension to each well of the black plate.
      3. Measure the fluorescence intensity at Ex. 540 nm and Em. 570 nm. For control, use wells containing unlabeled cells.
      Registered TrademarksCalbiochem® is a registered trademark of EMD Chemicals, Inc.
      Interactive Pathways™ is a trademark of EMD Chemicals, Inc.