I will perform MTT assays on cancer cells with 4 hours long treatments. Because of that, I would be finishing the MTTs too late, and my lab gets closed early. Can I leave the formazan crystals dissolving overnight so I can read it the next day?
Hello Osmar Rivera
Yes. You may leave the formazon crystals for dissolving overnight in the solubilization buffer at 37 degrees. It will not hamper your results.
For more insight, you can read the protocol given on MERCK website:
https://www.sigmaaldrich.com/IN/en/technical-documents/protocol/cell-culture-and-cell-culture-analysis/cell-counting-and-health-analysis/cell-proliferation-kit-i-mtt
Hello Osmar Rivera
The formazan crystals dissolve in solubilization solution within 15 to 20 minutes. Delaying the absorbance reading could be a problem as it could affect the stability of the formazan signal.
If you are using a commercial solubilization solution, then you would need to follow the instructions of the manufacturer who has optimized the conditions. If you are using for instance, acid-isopropanol to solubilize formazan crystals, the resultant color of the solubilized formazan has been reported to be stable for a few hours at room temperature.
So, it will depend on the solubilization solution that you use.
Best.
You can use SDS-HCl and leave the crystals dissolving overnight.
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