Cell Meter™ Fluorimetric Live Cell Cycle Assay Kit *Optimized for 405 nm Violet Laser Excitation*
Ordering information
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Additional ordering information
| Telephone | 1-800-990-8053 |
| Fax | 1-800-609-2943 |
| sales@aatbio.com | |
| International | See distributors |
| Bulk request | Inquire |
| Custom size | Inquire |
| Shipping | Standard overnight for United States, inquire for international |
Spectral properties
| Excitation (nm) | 401 |
| Emission (nm) | 460 |
Storage, safety and handling
| Intended use | Research Use Only (RUO) |
Alternative formats
| Cell Meter™ Fluorimetric Live Cell Cycle Assay Kit *Green Fluorescence Optimized for Flow Cytometry* |
| Cell Meter™ Fluorimetric Live Cell Cycle Assay Kit *Red Fluorescence Optimized for Flow Cytometry* |
Related products
| Overview |  SDSProtocol |
Excitation (nm) 401 | Emission (nm) 460 |
The cell cycle has four sequential phases: G0/G1, S, G2, and M. During a cell's passage through cell cycle, its DNA is duplicated in S (synthesis) phase and distributed equally between two daughter cells in M (mitosis) phase. These two phases are separated by two gap phases: G0/G1 and G2. The two gap phases provide time for the cell to grow and double the mass of their proteins and organelles. They are also used by the cells to monitor internal and external conditions before proceeding with the next phase of cell cycle. The cell's passage through cell cycle is controlled by a host of different regulatory proteins. This particular kit is designed to monitor cell cycle progression and proliferation by using our proprietary Nuclear Violet™ in live cells. The percentage of cells in a given sample that are in G0/G1, S and G2/M phases, as well as the cells in the sub-G1 phase prior to apoptosis can be determined by flow cytometry. Cells stained with Nuclear Violet™ can be monitored with a flow cytometer (Pacific Blue® channel).
Platform
Flow cytometer
| Excitation | 405 nm laser |
| Emission | 450/40 nm filter |
| Instrument specification(s) | Pacific Blue channel |
Components
Example protocol
AT A GLANCE
Protocol summary
- Prepare cells with test compounds at a density of 5 × 105 to 1 × 106 cells/mL
- Add 1 µL of 500X Nuclear Violet™ into 0.5 mL of cell solution
- Incubate at room temperature for 30 - 60 minutes
- Analyze with a flow cytometer using the 405 nm violet laser excitation
Important notes
Thaw all the components at room temperature before use.
SAMPLE EXPERIMENTAL PROTOCOL
- For each sample, prepare cells in 0.5 mL of 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.
- Treat cells with test compounds for a desired period of time to induce apoptosis or other cell cycle functions.
- Add 1 µL of 500X Nuclear Violet™ (Component A) into the cells containing the growth medium, and incubate the cells in a 37°C, 5% CO2 incubator for 30 to 60 minutes. 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 Nuclear Violet™. Note: The appropriate incubation time depends on the individual cell type and cell concentration used. Optimize the incubation time for each experiment. Note: It is not necessary to fix the cells before DNA staining since the Nuclear Violet™ is cell-permeable. Note: Human and rodent stem cells efflux Nuclear Violet™ stain, and this is the basis for the Side Population (SP) technique; the efflux can be blocked with blocking agents like verapamil, fumitermorgin C to prevent dye efflux for accurate DNA content analysis.
- Optional: Centrifuge the cells at 1000 rpm for 4 minutes, and then re-suspend cells in 0.5 mL of assay buffer (Component B) or the buffer of your choice.
- Monitor the fluorescence intensity by flow cytometry using the 405 nm violet laser (Ex/Em = 405/445 nm). Gate on the cells of interest, excluding debris.
Product Family
| Name | Excitation (nm) | Emission (nm) |
| Cell Meter™ Fluorimetric Fixed Cell Cycle Assay Kit *Red Fluorescence Optimized for Flow Cytometry* | 537 | 618 |
Images
Figure 1. DNA profile in growing Jurkat cells with Cell Meter™ Fluorimetric Cell Cycle Assay Kit. Jurkat cells were stained with Nuclear Violet™ for 30 minutes. The fluorescence intensity of Nuclear Violet™ was measured using ACEA NovoCyte flow cytometer with the channel of Pacific Blue. In growing Jurkat cells, G0/G1 and G2/M phase histogram peaks are separated by the S-phase distribution.
References
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