help! 293F cell line

Stephen Kwok Stephen.Kwok at tufts.edu
Fri Jan 18 20:44:57 EST 2008


Not sure if these cells are very fancy, but HEK 293 cells for 
tranfecting are very robust cells.  I used to culture these guys like 
mad in simple 10% DMEM media in a regular incubator at 7% C02.	If they 
are having troubles with debris, you might have a bacteria infection, 
or it simply does not like the SF media.

The cells hopefully are adapted to them since they are sending it to 
you, but I certainly wouldn't put it past that they were/are.

Try to bump it in some regular serum rich media and see if the growth 
is any better.	If it is, then you know they simply haven't been 
adapted.

Good luck, this shouldn't be too difficult at all.

Steve

Quoting Nancy Hurtado-Ziola <nhurtado at ucsd.edu>:

> Hello fellow flow-ers,
>
> I have a soluble protein that's really difficult to express.	I  
> mentioned it to someone and they said I should use 293F cells to  
> solve this problem.  Well, I need some information on these cells.   
> I'm having trouble growing 293F cells.	I purchased them directly  
> from Invitrogen.  The first batch started out OK.  I did everything  
> the supplier said to do (use their SF media, use warmed media, keep  
> cells at 37deg c, with 8% CO2 level, check viability with tripan  
> blue, etc., etc.)  but the cell population diminished and there was  
> so much debris.  I got to pass 3, but never had enough cells after  
> that to do another pass let alone freeze down or for a transfection.	
>  I've never seen a photo of what these cells are supposed to look 
> like	after the culture takes off.  Do cells in suspension typically 
> have a  lot of debris?
>
> I called Invitrogen and asked for help 3 different times during my  
> attempt to grow these cells.	They finally sent me a new vial of  
> cells.	Again, the cells started out ok.  The first pass, went ok,  
> but now the cells seem to be dying and there is an increase in
> cellular debris. (I also thought it might be the media so I gave them 
>  the lot number, but there didn't seem to be any reported problems.)	
>  Has ANYONE seen this before?  If so, did you figure out what the  
> problem was?	Were you able to resolve it and get a good culture 
> going?
>
> As always, I appreciate any help you might have to offer.
> ________________________
> Nancy Hurtado-Ziola, Ph.D.
> Varki Laboratory
>
> Department of Cellular and Molecular Medicine
> University of California, San Diego
> 9500 Gilman Dr.
> CMM-E 1087, Mail Code 0687
> La Jolla, CA 92093
> Lab Phone: (858) 534-1346
> Fax: (858) 534-5611
>
>






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