Cell-mediated Cytotoxicity Assay: Basic Cytotoxicity Test
The Basic Cytotoxicity Test Assay Kit is a single-tube, dual-color assay for determining cell toxicity by flow cytometry. The assay employs a green fluorescent cellular stain, CFSE, to label target cells and the red live/dead viability dye 7-AAD to identify the dead cells present in the cytotoxicity assay samples. Analyze your results using a flow cytometer.
Cell-mediated Cytotoxicity Assay: Basic Cytotoxicity Test
Cytolytic activity is an important process for eliminated intracellular pathogens and cancer cells. ICT’s Basic Cytotoxicity Assay Kit allows you to assess cytolytic activity in cell culture.
The Basic Cytotoxicity Assay Kit includes two fluorescent reagents: CFSE and 7-AAD. CFSE, a green fluorescing membrane stain, is utilized to identify the target cell population. The unstained effector cells are added and incubated with the target cells. 7-AAD, a red fluorescing live/dead stain, is then added to stain all necrotic cells red by binding to the DNA of membrane-compromised cells. The target cell and effector cell populations can then be easily distinguished, and flow cytometry easily identifies live and necrotic cells.
7-AAD, CFSE
Cell-mediated cytotoxicity
7-AAD – 546 nm / 647 nm
CFSE – 492 nm / 520 nm
Flow cytometry
Cell culture
CFSE at ≤ -20°C, other components at 2-8°C
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United States
- Prepare samples and controls
- Dilute Assay Buffer 1:10 with diH2O and filter sterilize.
- Reconstitute CFSE with 200 µL DMSO and dilute 1:250 with 1X Assay Buffer.
- Add 200 µL CFSE to target cells to stain them green and incubate 15 minutes at room temperature.
- Wash target cells and adjust cell concentration.
- Add effector cells to target cells.
- Reconstitute 7-AAD with 260 µL DMSO and dilute 1:10 with 1X Assay Buffer.
- Add 20 µL 7-AAD to label necrotic cells red and incubate 10 minutes on ice.
- Run controls and analyze with a flow cytometer. Identify target cells by analyzing FL-1 (green) vs. FL-3 (red). CFSE excites at 492 nm and emits at 520-540 nm; 7-AAD excites at 546 nm and emits at 647 nm.
Kit 969: 125 Tests
CFSE target cell stain, 1 vial, #6162
7-AAD vital stain, 1 vial, #6163
10X Assay Buffer, 30 mL, #6161
Kit Manual
Kit 970: 250 Tests
CFSE target cell stain, 1 vial, #6162
7-AAD vital stain, 2 vials, #6163
10X Assay Buffer, 60 mL, #685
Kit Manual
Product Specific References
| PMID | Publication |
| 37120009 | Lee, M.H., et al. 2023. Live Mycobacterium paragordonae induces heterologous immunity of natural killer cells by eliciting type I interferons from dendritic cells via STING-dependent sensing of cyclic-di-GMP. Microbes and infection, 105144. |