logo
AAT Bioquest

Cell Meter™ Caspase 3/7 Activity Apoptosis Assay Kit *Green Fluorescence Optimized for Flow Cytometry*

Our Cell Meter™ assay kits are a set of tools for monitoring cell viability. There are a variety of parameters that can be used for monitoring cell viability. This particular kit is designed to monitor cell apoptosis by measuring generic caspase 3/7 activation in living cells. Caspase 3/7 activation is widely accepted as a reliable indicator for cell apoptosis. Caspases have substrate selectivity for the peptide sequence Asp-Glu-Val-Asp (DEVD). This kit uses TF2-DEVD-FMK as a fluorogenic indicator for caspase 3/7 activity. TF2-DEVD-FMK is cell permeable, nontoxic, and irreversibly binds to activated caspase 3/7 in apoptotic cells. Once bound to caspases, the green fluorescent reagent is retained within the cell. The binding event prevents the caspases from further catalysis but will not stop apoptosis from proceeding. The reagent will start to react with active caspase enzymes within 15 minutes of addition to the media. The kit provides all the essential components with an optimized assay protocol. It is used for the quantification of most activated caspase activities in apoptotic cells, or for screening caspase inhibitors. The green label allows for direct detection of activated caspases in apoptotic cells by flow cytometry.

Example protocol

AT A GLANCE

Protocol summary

  1. Prepare cells with test compounds at a density of 5 × 105 to 1 × 106 cells/mL
  2. Add 1 µL of 500X TF2-DEVD-FMK into 0.5 mL of cell solution
  3. Incubate at 37°C, 5% CO2 incubator for 1 - 4 hours
  4. Pellet the cells and resuspend the cells in 0.5 mL of assay buffer or growth medium
  5. Analyze cells using flow cytometer with 530/30 nm filter (FITC channel)

Important notes
Thaw all the components at room temperature before starting the experiment.

SAMPLE EXPERIMENTAL PROTOCOL

  1. For each sample, prepare cells in 0.5 mL warm medium or buffer of your choice at a density of 5×105 to 1×106 cells/mL. Note: Each cell line should be evaluated on an individual basis to determine the optimal cell density for apoptosis induction.

  2. Treat cells with test compounds for a desired period of time to induce apoptosis, and create positive and negative controls.

  3. Add 1 µL of 500X TF2-DEVD-FMK (Component A) into the treated cells.

  4. Incubate the cells in a 37°C, 5% CO2 incubator for 1 - 4 hours. Note: For adherent cells, gently lift the cells with 0.5 mM EDTA to keep the cells intact, and wash the cells once with serum-containing media prior to incubation with TF2-DEVD-FMK. The appropriate incubation time depends on the individual cell type and cell concentration used. Optimize the incubation time for each experiment.

  5. Wash and spin the cells twice. Resuspend the cells in 0.5 mL of assay buffer or growth medium. Note: TF2-DEVD-FMK is fluorescent; theforefore it is important to wash out any unbound reagent to remove the background.

  6. If desired, label the cells with a DNA stain (such as propidium iodide or 7-AAD for dead cells).

  7. If desired, fix cells.

  8. Monitor the fluorescence intensity using a flow cytometer with 530/30 nm filter (FITC channel). Gate on the cells of interest, excluding debris.

Spectrum

Citations

View all 38 citations: Citation Explorer
High-dose ascorbate exerts anti-tumor activities and improves inhibitory effect of carboplatin through the pro-oxidant function pathway in uterine serous carcinoma cell lines
Authors: Shen, Xiaochang and Wang, Jiandong and Deng, Boer and Chen, Shuning and John, Catherine and Zhao, Ziyi and Sinha, Nikita and Haag, Jennifer and Sun, Wenchuan and Kong, Weimin and others,
Journal: Gynecologic Oncology (2024): 93--102
Hexamethylene Amiloride synergizes with Venetoclax to induce lysosome-dependent cell death in acute myeloid leukemia
Authors: Jiang, Xinya and Huang, Kexiu and Sun, Xiaofan and Li, Yue and Hua, Lei and Liu, Fangshu and Huang, Rui and Du, Juan and Zeng, Hui
Journal: iScience (2023)
Metformin sensitizes AML cells to venetoclax through endoplasmic reticulum stress—CHOP pathway
Authors: Hua, Lei and Yang, Nianhui and Li, Yue and Huang, Kexiu and Jiang, Xinya and Liu, Fangshu and Yu, Zhi and Chen, Jie and Lai, Jing and Du, Juan and others,
Journal: British Journal of Haematology (2023)
GONIOTHALAMIN INHIBITS CELL GROWTH, PERTURBS CELL CYCLE AND INDUCES APOPTOSIS IN HUMAN OSTEOSARCOMA SAOS-2 CELLS: Received 2023-01-03; Accepted 2023-02-15; Published 2023-06-06
Authors: Bakar, Siti Aishah Abu and Azhar, Nur Asna and Mohamad, Noor Muzamil and Nordin, Ira Maya Sophia and Ali, Abdul Manaf and Noor, Siti Noor Fazliah Mohd and Hamid, Shahrul Bariyah Sahul and Ahmad, Nor Hazwani
Journal: Journal of Health and Translational Medicine (JUMMEC) (2023): 105--115
Page updated on December 6, 2024

Ordering information

Price
Unit size
Catalog Number22823
Quantity
Add to cart

Additional ordering information

Telephone1-800-990-8053
Fax1-800-609-2943
Emailsales@aatbio.com
InternationalSee distributors
Bulk requestInquire
Custom sizeInquire
Technical SupportContact us
Purchase orderSend to sales@aatbio.com
ShippingStandard overnight for United States, inquire for international
Request quotation

Spectral properties

Correction Factor (280 nm)

0.09

Extinction coefficient (cm -1 M -1)

75000

Excitation (nm)

503

Emission (nm)

525

Quantum yield

0.91

Storage, safety and handling

Certificate of OriginDownload PDF
H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22
UNSPSC12352200

Platform

Flow cytometer

Excitation488 nm laser
Emission530, 30 nm filter
Instrument specification(s)FITC channel

Components

Detection of caspase 3/7 activities using Cell Meter™ Caspase 3/7 Activity Apoptosis Assay Kit in Jurkat cells. TF2-DEVD-FMK fluorescence intensity was induced with the addition of camptothecin. Jurkat cells were treated without (Blue) or with 20 µM camptothecin (Red) in a 37 °C, 5% CO2 incubator for 4-5 hours, and then dye loaded with TF2-DEVD-FMK for 1 hour. Response was measured using BD FACSCalibur using FL1 channel.
Detection of caspase 3/7 activities using Cell Meter™ Caspase 3/7 Activity Apoptosis Assay Kit in Jurkat cells. TF2-DEVD-FMK fluorescence intensity was induced with the addition of camptothecin. Jurkat cells were treated without (Blue) or with 20 µM camptothecin (Red) in a 37 °C, 5% CO2 incubator for 4-5 hours, and then dye loaded with TF2-DEVD-FMK for 1 hour. Response was measured using BD FACSCalibur using FL1 channel.
Detection of caspase 3/7 activities using Cell Meter™ Caspase 3/7 Activity Apoptosis Assay Kit in Jurkat cells. TF2-DEVD-FMK fluorescence intensity was induced with the addition of camptothecin. Jurkat cells were treated without (Blue) or with 20 µM camptothecin (Red) in a 37 °C, 5% CO2 incubator for 4-5 hours, and then dye loaded with TF2-DEVD-FMK for 1 hour. Response was measured using BD FACSCalibur using FL1 channel.