I have trouble inducing autophagy in HeLa Cell. HeLa cell was transfected with pmCherry-LC3B for 24 hours, and EBSS（with 0.1% w/v glucose) was used to induce autophagy for 2 hours.The IF image is showed in the attachment.There are small portion of puncta positive cells in my specimen. As I have read some papers showing much more number of LC3B puncta structure in cytoplasm during autophagy triggered by HBSS or EBSS for 2 hours,so I just wonder whether there are some key point in this experiment?
Sorry everybody who has been visiting this question.I think I have found the reason.I have a double check on my sequencing result,and to my surprise, the sequence is quite different to the one I put into BLAST.Maybe I have open too much files and copy the template sequence for BLAST by mistake that day when I was checking this construct.
That's right. There was no other reason to do not being able to induce autophagy to Hela cells, since your experimental design is great. I myself have a paper inducing a toxin induced vacuolating efect on Vero, Hela and other cells, that turned out later on to be autophagy. Good luck with experiments.
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