QIA90 Caspase Detection Kit (FITC-VAD-FMK)

QIA90
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Overview

Replacement Information

Key Specifications Table

Detection Methods
Fluorescence

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QIA90-1KIT
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      Description
      OverviewA convenient and sensitive method for the detection of activated caspases in living cells. The fluorescent marker, FITC-VAD-FMK, is a cell-permeable, non-toxic inhibitor that binds irreversibly to activated caspases in apoptotic cells. The fluorescence intensity can be measured by fluorescence microscopy, fluorescence plate reader, or flow cytometry.
      Catalogue NumberQIA90
      Brand Family Calbiochem®
      References
      Product Information
      Detection method Fluorescence
      Form100 Tests
      FormatFlow Cytometry or Fluorimeter
      Kit containsLabeled Caspase Inhibitor (FITC-VAD-FMK), Wash Buffer, 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
      R PhraseR: 36/37/38

      Irritating to eyes, respiratory system and skin.
      S PhraseS: 23-26-38

      Do not breathe fumes.
      In case of contact with eyes, rinse immediately with plenty of water and seek medical advice.
      In case of insufficient ventilation, wear suitable respiratory equipment.
      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.
      Avoid freeze/thaw Avoid freeze/thaw
      Do not freeze Ok to freeze
      Packaging Information
      Transport Information
      Supplemental Information
      Kit containsLabeled Caspase Inhibitor (FITC-VAD-FMK), Wash Buffer, Caspase Inhibitor (Z-VAD-FMK), and a user protocol.
      Specifications

      Documentation

      Caspase Detection Kit (FITC-VAD-FMK) SDS

      Title

      Safety Data Sheet (SDS) 

      Caspase Detection Kit (FITC-VAD-FMK) Certificates of Analysis

      TitleLot Number
      QIA90

      Brochure

      Title
      The Active Siteル Volume 4, Number 1
      User Protocol

      Revision10-August-2007 RFH
      Form100 Tests
      FormatFlow Cytometry or Fluorimeter
      Detection method Fluorescence
      Speciesa broad range of species
      StorageUpon arrival store the entire contents of the kit at -20°C.
      Principles of the assayThe Calbiochem® Caspase Detection Kit (FITC-VAD-FMK) provides a convenient and sensitive means for detecting activated caspases in situ in living cells. The assay utilizes a caspase inhibitor (VAD-FMK) conjugated to FITC as the fluorescent in situ marker. FITC-VAD-FMK is cell permeable and nontoxic and irreversibly binds to activated caspases in apoptotic cells. The FITC label allows for direct detection of activated caspases in apoptotic cells by fluorescence microscopy, flow cytometry, or fluorescence plate reader.
      Materials provided• FITC-VAD-FMK (Kit Component No. JA5600): 1 vial, 100 µl each
      • Wash Buffer (Kit Component No. JA5601): 2 bottles 100 ml each
      • Z-VAD-FMK (Kit Component No. JA5602): 1 vial, 10 µl
      Detailed protocolA. Staining Protocol:
      1. Induce apoptosis in cells (1 x 106/ml) by desired method. Concurrently incubate a control culture without induction. An additional control can be prepared by adding the caspase inhibitor Z-VAD-FMK at 1 µl/ml culture to an induced culture to inhibit caspase activity.
      2. Aliquot 300 µl each of the induced and control cultures into microfuge tubes.
      3. Add 1 µl FITC-VAD-FMK into each tube and incubate for 0.5-1 h in a 37°C incubator with 5% CO2.
      4. Centrifuge the cells at 3000 rpm for 5 min and discard supernatant.
      5. Resuspend the cells in 0.5 ml Wash Buffer and centrifuge as above. Repeat for a total of 2 washes.
      6. Proceed to analysis using one of the methods below.

      B. Quantification by Flow Cytometry
      For flow cytometric analysis, resuspend the cells in 300 µl Wash Buffer. Put samples on ice. Analyze samples by flow cytometry using the FL-1 channel.

      C. Detection by Fluorescence Microscopy
      For fluorescence microscopy, resuspend the cells in 100 µl Wash Buffer. Put one drop of the cell suspension onto a slide and cover with a coverslip. Observe cells under the fluorescence microscope using a FITC filter. Caspase positive cells appear to have brighter green signals, whereas caspase negative control cells show much weaker signal.

      D. Analysis by Fluorescence Plate Reader
      For analysis with a fluorescence plate reader, resuspend the cells in 100 µl Wash Buffer and transfer the cell suspension to designated wells of the black plate. Measure the fluorescence intensity at an Exitation of ~485 nm and an Emission of = ~535 nm. For a 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.