I am doing experiments to determine the cytokine response of HD11 cells after infection with bacteria....if, during my experiment, I use trypsin to remove my cells, will this impact my ability to extract RNA from the cells?
Thanks!
It is possible that the detachment of the cells will start a program within the cells that change the expression of certain genes but the process of RNA preparation should not be disturbed. Most kits use a lysis buffer as a first step and we never saw an interference when using the cell pellet after trypsin detachment for RNA preparation. On the other hand we generally use the lysis buffer directly on the plate without trypsin treatment.
Yes - many (animal-derived) trypsin reagents should be avoided and there was a recent important (2014) paper on avoiding the use of trypsin for this purpose at: http://www.ncbi.nlm.nih.gov/pubmed/24983903
RNAse content of certain trypsin reagents is the danger.
Michael Marcus Hoffman's above suggestion of putting the lysis reagent directly on the cells is a good way out of this.
Thank you, I know a number of people that use trypsin and do not have issues, but it concerned me that it may not be the best thing to use.
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