According to my protocol i have to incubate my plate for 1 hr after adding MTT solution. but i incubated for 2.45 hours. My results were not good. Some variations in the OD values. My cell density was good in each well when i seeded cells to my wells on 1st day, Just want to know whether long incubation time with MTT affects the OD values?
Sonia, there should be no issues leaving cells in MTT solution. It is not unusual to do so for 3 hours with some cell lines (e.g. those with low levels of succinate reductase).
Hope this helps
Jon
Sonia- since you are measuring metabolism rather than proliferation and different adeoncarcinoma cell lines vary in metabolic rate depending on the mutations,- you might try optimizing the MTT assay for each different adenocarinoma cell line i.e different amounts of time in the MTT solution, cell density etc. Start with one cell line only.
Sonia- As mentioned by John, you may vary the number of cells per well. It seems that either your cells are highly metabolically active or you may have seeded more cells.
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