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Why are my Huh 7s dying after transfections?

Huh 7s tissue culture transfection

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#1 KhaleesiDany

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Posted 06 October 2015 - 11:27 PM

Greetings

 

I have been trying to transfect my Huh 7 cells, but each time after 48 hours from transfection in the centre of well i see small rounded cells they not even floating but i think they dead. I can only see gfp on the outskirts of well where the cells are not tiny. I tried PEI, lipofectamine and jet prime, they still die, i always do transfections around 3pm- 4pm and change media the next morning. I use antibiotic containing media, but with lipofectamine i use antibiotic free media. Only once did i find that my cells never died so i continued to do immunofluoresnce staining but i couldnt pick up any signal from antibodies, the GFP control wells showed that transfection did work, i did the same procedure with hek 293 cells and i could pick up antibody signal so the problem is not the antibodies any suggestion on why my cells are dying? 



#2 gvbdxz

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Posted 09 October 2015 - 02:32 AM

I haven't worked with these cells but clearly, you're leaving your cells with the tranfection agent (PEI, lipofectamine and jet prime) for much too long. Try changing your medium after 4 hours or transfection, no more. It'll reduce the toxicity. 

 

Hope it helps!


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