MTT assay medium with phenol red - (Mar/22/2011 )

Can MTT assay be performed in medium with phenol red?

Make 5mg/mL MTT dissolved in PBS
5 hours before the end of incubation add 20uL of MTT solution to the wells containing cells (cells in medium with phenol red)
incubation 5h 37C
removing media
Adding 200uL of DMSO to dissolve cristals,
Reading 550nm

kamilaf on Tue Mar 22 11:47:00 2011 said:

That's exactly how we perform MTT assay in our lab. The only difference is that we incubate the cells for 4 instead of 5 hours. There is no problem with the phenol red. As you mentioned the mediun is thrown away before DMSO addition and the colour you finally take in your wells comes only from the crystals formed inside mitochondria of living cells.
So you don't have to worry about Phenol Red at all!

Hi, thanks for answer, I was doing also 4h incubation, only I am not sure about reading, I tried in 550nm, and reference 620, but I am not sure if I need to use this reference or not?
Generally it looks quite good!

Cheers!