Both non-adherent cells and adherent cells are suitable. All operations are performed under sterile conditions. Avoid contaminating cells, because microbial contamination will also reduce alamarBlueTM, antibiotics such as penicillin G (1U / ml), amphotericin B (amphotericin B, 0.0025ug / ml) should be added to the medium.
The redox indicator is pH sensitive, and phosphate buffer is recommended. 10% fetal calf serum has no effect on colorimetric detection, but it will quench part of the fluorescence; therefore, 10% BSA needs to be added to the control solution when using fluorescence detection. Phenol red also has a certain influence on the detection, and the result will be increased by 0.03%. In general, non-reducing media should be used, such as RPMI1640, Hank's-Eagle or Dulbecco's-Eagle.
When used in a special system, the conditions such as cell density, incubation time, medium composition, and use concentration need to be optimized first.
Collect the cells to be tested and plate them on fresh medium supplemented with 10% (v / v) alamarBlueTM at an optimized density.
1. Cell culture: RPMI medium, add HEPES, 10% fetal bovine serum, 1% glutamate, appropriate antibiotics;
2. When the cell coverage reaches 80%, use trypsin digestion and spread it in a microplate with a density of 825 ~ 80,000 cells per well.
After 3.1 hours, add alamarBlueTM and incubate at 37 ° C with 5% CO2
4. Check the reduction of alamarBlueTM after the scheduled time point , An anticancer drug, cyclin kinase inhibitor), trypsin digest the cells, collect and spread on the medium containing the drug, and culture at 37 ° C with 5% CO2 for 2 days; the medium before removal is replaced with 10% ( v / v) New culture medium of alamarBlueTM, and check the reduction degree at predetermined time points.
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