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Changing the media after transfection of 293T cells - I did not change the media the day after transfecting 293T cells. (Aug/16/2010 )

So I transfected 293T cells with 5 different plasmids on Saturday. I was supposed to change the media on Sunday, but the door to the tissue culture room was locked and I could not get in.

Do you think the cells will still be OK and the transfection will work? Thank you!

-distalless-

distalless on Mon Aug 16 17:02:18 2010 said:


So I transfected 293T cells with 5 different plasmids on Saturday. I was supposed to change the media on Sunday, but the door to the tissue culture room was locked and I could not get in.

Do you think the cells will still be OK and the transfection will work? Thank you!


293t are quite tough, so if they are still alive ( I dont know what kind of reagent did you use, and what is the toxicity) I would continue with the experiment

-laurequillo-

laurequillo on Mon Aug 16 17:07:24 2010 said:


distalless on Mon Aug 16 17:02:18 2010 said:


So I transfected 293T cells with 5 different plasmids on Saturday. I was supposed to change the media on Sunday, but the door to the tissue culture room was locked and I could not get in.

Do you think the cells will still be OK and the transfection will work? Thank you!


293t are quite tough, so if they are still alive ( I dont know what kind of reagent did you use, and what is the toxicity) I would continue with the experiment


I used Lipofectamine 2000. How would I be able to tell if they are still alive? The plate seems very confluent.

-distalless-

If the plate is confluent, the cells are probably alive - dead cells are usually rounded and floating.

-bob1-

bob1 on Mon Aug 16 23:17:34 2010 said:


If the plate is confluent, the cells are probably alive - dead cells are usually rounded and floating.


Yes the plate is confluent, but the cells are coming off the plate. In fact more than half of the cells have peeled off the plate!

-distalless-

that's probably because your 293T's are over-confluent...not because of the transfectionmix

-fysio lab-

fysio lab on Tue Aug 17 08:11:03 2010 said:


that's probably because your 293T's are over-confluent...not because of the transfectionmix


Do you think that I can still go ahead and use the supernatant for viral transduction? Thanks!

-distalless-

distalless on Tue Aug 17 18:38:48 2010 said:


fysio lab on Tue Aug 17 08:11:03 2010 said:


that's probably because your 293T's are over-confluent...not because of the transfectionmix


Do you think that I can still go ahead and use the supernatant for viral transduction? Thanks!


i think so, but pass the medium over a 0.45 filter to get rid of the dead cells

-fysio lab-