Adding a drug causes cells to undergo oxidative stress as indicated by DCFDA fluorescence and an increase of ABCG2. However, I get inconsistent results of SOD1 after 24 hours of exposure. Also, if you want to get the expression, do you collect only the remaining live cells or also collect the floating/apoptotic cells?
I'm not sure I'm on the same theme as you, but I've been able to work on oxidative stress in aquatic organisms.
But from an external point of view, if you take the apoptotic and/or floating cells, you will introduce a bias in the analysis because these cells seem already dead and therefore non-functional. They are probably already degrading and will not be representative of what is happening in your experiment.
As SOD1 is generally treated as "constitutive" then it is really the tricky one. https://pubmed.ncbi.nlm.nih.gov/25908445/
Eric Gismondi: yes, I do agree. I compared both conditions and found out that taking the whole population (with the apoptotic cells) diluted the protein expression, thus making it inaccurate.
Adam Figarski: I see. Thank you for the reference. In your opinion, could it be possible that the cells undergoing oxidative stress would increase their gene transcription as a physiological reaction but, to the downstream effects, the protein SOD1 expression would be decreased as it will be being used out to convert all O2-?
Read this article
https://www.sciencedirect.com/science/article/pii/S2213231717302574#:~:text=Increased%20expression%20of%20SOD1%20may,9%5D%2C%20%5B13%5D.
Functioning of Superoxide Dismutase is increased, because generation of free radicals produce adverse effects. SOD catalyze the chemical reactions having single oxygen and slow down or prevent the generation of free radicals. So, automatically SOD activity increased...
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