Dear Mude Hemanjali,
unfortunately you added no additional information concerning your assay..!
One explanation for your observation could be that control cells simply have overgrown and started to detach (did the medium already turn yellow)? If treatment is growth limiting for example, cells in those treated wells would still give a signal with the MTT assay.
Maybe try seeding less cells or shorten the incubation time?
Hope that helps,
best regards,
Christian
As there is very minimal information in your question, its not possible a straight forward answer without asking you multiple questions !
1. What cells are being tested? are they adherent type? If not, it's not a surprise that you don't see a signal ! On the other hand, the treatments may be inducing adhesion in the cells. Just a guess ! Plate bases MTT assays are not the right choice for non-adherent cells.
2. Thinking that the cells are adherent type, next possibilities to think about are whether or not cells in the control wells are added with MTT + solubilizing buffer (10% SDS in 0.1 N HCL) !
3. Are you seeing no signal in negative control or in blank? Negative control for MTT is untreated cells + MTT + solubilizing buffer while blank is medium with NO cell + MTT + Solubilizing buffer !!
BW
In my opinion, you should check and maybe revise the experimental protocol/setting. Adherence of cells in not necessary, you can also check cell viability via MTT assays with non-adherent cells. However, if you want to test adherent cells, you should avoid loss (e.g. via washing steps etc.). This could be reached via usage of adequate plastics/plates, coating etc. However, this depends on your experimental setting, as mentioned before.
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