# Population doubling time of keratinocyte,etc. - (Mar/01/2005 )

again thanks!!

i've done some preliminary tests, what i suspected was the period of keratinocyte adherent might be due to some factors. around the results i have got was the seeding density. if i seed too less cells, cell could adhere but need long period to get 60% of confluency and of course some cells were proliferated and DIFFERENTIATED. this is worse!

if too much cells, medium will need to change everyday, but 80% confluency can be achieved within 4 days. donor variability also a factor in this aspect as well.

what do u think about the seeding density??

thanks for comments!!

hi

you must pay attention to the time cells need to differenciate. if your cells need 5 days to differenciate, i would say that three days is the max time for your experiment. But if time of differenciation is greater no problem fo doing your experiment on 4 days!

I think that 60% confluency is good. For two reasons. 1 cells are relatively close to make good divisions (but not too close) and 2 if you seed at more than 60% and due to the fact your treatment is an increaser of division frequency (PDT decreases) more than 60% may result in too much confluency in treated cells.

Enjoy your experiment. Good luck.

Fred.

thanks,fred!!

u are right. i'll try through out the passage 0 to 2 with treatment, to observe the differences of PDT among passage 0,1 and 2. what i guess was the passage 2 will be cleaner than the other two. thus, will be giving out the more accurate answer. what do u think??

thanks!!

Hi,

I am stimating PDT for a normal and transformed cell line but I cannot find the formula to calculate it. Does any one has this formula? I am plating 50,000 or 100,000 cells in 35mm dishes and counting cells 24, 48, and 72 hours later.

I will appreciate your help. Thanks.

David

here you are...

negative sign may occur but you don't care of it...

Thanks for the formula for PDT.

As I can see, I can use only two time points at a time, is that correct? I have data collected at times 0, 24, 48, and 72 hours.

Should I compared PDT between 0-24, 0-48, and 0-72, using t=24, 48, and 72 respectively, then average? or should I compara 0-24, 24-48, and 48-72 using t=24 for all, then average?

How would you use the formula to obtain your cell line's PTD?

Thanks

Can anyone send me keratinocyte isolation protocol?

Thanx U