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Splitting Cells that are shaped as small spheres

splitting cell culture

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#1 procrasemit

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Posted 23 July 2014 - 08:21 AM

I'm currently growing FRTL-5 cells (rat thyroid cell line). We have recently encountered a problem with these cells after having transfected them. Whereas before they used to be growing in a nice monolayer (prior to transfection), they are now in the forms of clustered spheres. 

 

Would it still be possible for us to detach these cells and re-plate them again. If so, are there any necessary precautions that we should take into account before doing so? Thanks in advance for the help! :)



#2 bob1

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Posted 23 July 2014 - 07:23 PM

By spheres do you mean floating clusters of cells or attached clumps?

 

It should be possible to split and re-plate, but - as you might have noticed - the transfection has altered the behaviour of your cells in a manner that seems to have affected their attachment, which might mean that they won't take well to splitting.

 

Basically, the only thing to do is try it and see what happens. It would probably pay to try other methods of splitting too.



#3 SusieQ

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Posted 28 July 2014 - 10:28 PM

I have some experience with cells that cluster during culturing. What I do to get them to detach from each other is, after trypsinization and adding fresh culture medium, pipetting up and down with a 10 ml volume pipette with a 200 ul pipet tip attached to it. Basically, I pipet up the cells and then pipet them back in by putting the tip of the pipet in the corner of a flask, so that the cells are forced to go through a very small opening. With my PC12 cells it works pretty good, and those cells are sometimes a pain to detach. However, keep in mind that this method delivers a lot of mechanical stress on the cells and I believe not every cell line can survive this method. 

 

Like bob1 said, just try it out while maintaining sufficient back-up material in case things go bad.







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