cell confluency & effect on cells...?? - (Aug/03/2010 )
Hi Everyone,
I was trying to findout if splitting cells continuously at 50 - 60% confluency or splitting them when they are over confluent bring any changes to the cell line.
if they dont why exactly do we split cells between 80-95% confluency...?? please help understand the reason behind this.
thanks in advance
k.sagi
With any cell line the aim is to make sure that the cells are not undergoing selective processes that may change the phenotype of the cells... splitting them when they are at too low a density will cause selection for cells that can survive well at low densities. Leaving them to become over confluent results in cell populations that are able to function well at high density. Either of these states may be "normal" for the cell line, but if you do these sorts of things it is a good idea to compare them to the parent line every so often to ensure that they haven't changed behavior, morphology etc.
This is the reason why publications now often ask for proof that the cell lines you are using are validated to be the cell line that you said they are.
bob1 on Tue Aug 3 22:18:46 2010 said:
With any cell line the aim is to make sure that the cells are not undergoing selective processes that may change the phenotype of the cells... splitting them when they are at too low a density will cause selection for cells that can survive well at low densities. Leaving them to become over confluent results in cell populations that are able to function well at high density. Either of these states may be "normal" for the cell line, but if you do these sorts of things it is a good idea to compare them to the parent line every so often to ensure that they haven't changed behavior, morphology etc.
This is the reason why publications now often ask for proof that the cell lines you are using are validated to be the cell line that you said they are.
Hi there,
thanks for the reply thats really helpful. regarding the comparision of the cell lines how do we do it..?
is it using methods like isoenzyme analysis and PCR or something else. i also wanted to ask if you end up selecting the cells would that affect your assay in way. (like if you have selected cells that grow well at low densities would they still survive at higher densities and things like that). i know this sounds like a stupid question but i really want to what happens...??
cheers
k.sagi
Characterisation by isozymes may work, a lot of characteriation is done based on growth rates, morphology, and responses to particular treatments. Basically you can compare your cells with new ones for your system to ensure that there are no significant changes in your model. Selection pressures may or may not affect how the cells behave during your experiments.
Also, if you passage at low density you start to select for faster growing cells over time--if you let them get too confluent you start changing the gene expression patterns due to contact and nutrient inhibition. In either case, you start to change the cell line like Bob1 said.