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QIA113 Caspase-8 Detection Kit (FITC-IETD-FMK)

Caspase-8 Detection Kit (FITC-IETD-FMK) MSDS (material safety data sheet) or SDS, CoA and CoQ, dossiers, brochures and other available documents.
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QIA113
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Overview

Replacement Information

Key Spec Table

Detection Methods
Fluorescence

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      Description
      OverviewA fast, convenient, and sensitive method to measure caspase-8 in living cells. The fluorescent marker, FITC-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 in a fluorescent microplate reader, flow cytometer, or by fluorescence microscopy.
      Catalogue NumberQIA113
      Brand Family Calbiochem®
      References
      Product Information
      Detection methodFluorescence
      Form100 Tests
      FormatFlow cytometry or fluorescence microscopy
      Kit containsLabeled Caspase-8 Inhibitor (FITC-IETD-FMK), Wash Buffer, Unlabeled Caspase Inhibitor (Z-VAD-FMK), and a user protocol.
      Quality LevelMQ100
      Applications
      Biological Information
      Assay time1.5 h
      Sample TypeIntact cells
      Species Reactivity
      • A Broad Range Of Species
      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 (FITC-IETD-FMK), Wash Buffer, Unlabeled Caspase Inhibitor (Z-VAD-FMK), and a user protocol.
      Specifications
      Global Trade Item Number
      Catalogue Number GTIN
      QIA113-1KIT 04055977209358

      Documentation

      Caspase-8 Detection Kit (FITC-IETD-FMK) SDS

      Title

      Safety Data Sheet (SDS) 

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

      TitleLot Number
      QIA113
      User Protocol

      Revision02-Febuary-2009 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 assayThis Calbiochem® brand Caspase-8 Detection Kit provides a convenient and sensitive means for detecting activated caspase-8 in living cells. The assay employs a Caspase-8 inhibitor conjugated to FITC, the fluorescent marker FITC-IETD-FMK, is a cell-permeable, non-toxic inhibitor that binds irreversibly to activated caspase-8 and allows the detection of Caspase-8 by the measurement of fluorescence intensity (excitation max.: ~485 nm; emission max.: ~535 nm) in a fluorescent microplate reader, by flow cytometry, or by fluorescence microscopy.
      Materials provided• FITC-IETD-FMK (Kit Component No. JA7620-100UL): 1 vial, 100 µl
      • Wash Buffer (Kit Component No. JA7621-100ML): 2 bottles, 100 ml
      • Z-VAD-FMK (Kit Component No. JA7622-10UL): 1 vial, 10 µl
      Detailed protocolCaspase-8 Assay Procedures

      Staining

      1. Induce apoptosis in cells (1 x 106/ml) as desired. Concurrently incubate a control culture without apoptosis 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 activity.
      2. Aliquot 300 µl each of the induced and control cultures into eppendorf tubes.
      3. Add 1 µl of FITC-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 the supernatant.
      5. Resuspend cells in 0.5 ml of wash buffer and centrifuge again at 3000 rpm for 5 min.
      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-1 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 FITC filter. Caspase-8 positive cells appear to have brighter green signals, whereas caspase-8 negative control cells show a 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. max. 485 nm and Em. max. 535 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.