I'm currently testing the antiproliferative effect of drug on a suspension cell line called (Raji) and seems to be very easy to handle and grow. i have been seeding them into a regular 96-well plate according to one article i read. with a density of 5000 cells in a total 200 ul/well medium so i don't have to change medium before treatment and not to cause any cell loss.
can you please advise me on whether my technique is correct or not? if not what is the right equipment for seeding a suspension cell line?
Dear Enas,
I think this can theoretically be done, but maybe it should be carried out in a 24 wells, in order to provide cells with enough space and nutritive factors. Otherwise, cells could die due to confluence state.
Good luck with your work,
Davide
i often use this technique and i reach good resuslts. You should Pay attention on cell number for each well and on the confluence state.
We regularly plate lymphoid cell lines (SupT1, MT4) as well as monocytic cultures (U-937) in 96-well plates for pseudotype infection, no problems encountered. What do you mean by "no filter"?
Dear Enas,
the required time for cell assay must be taken in to account. If the time you need is so high that the color of the medium changed, the cells may die due to nutrition shortage.
you can reduce the volume to 100ul and increase the cell count up to 10,000 per well. low volume will decrease the pressure on cells at the bottom of the well.
I regularly perform this assay in 96 well plates (flat bottom wells) on both suspension and adherent cell lines. I always use 100ul/well and most often 10,000 cells/well. Just make sure to have control wells (cells only, no drug), and set these to 100%.
Do take into account the required time for the cell assay. Most often you may find this in other publications, yet sometimes you may have to assess this yourself: Think how the drug works ie how long until you would see the desired effect, and then keep this the same throughout.
Good luck!
Dear Prof. Zaheen,
Thank you so much for your answer. If i reduce the volume to a final 100uL/ well I will disturb the cell count because of the nature of the suspension cell. so what i do,I seed the cells in 100 uL then after 24 hrs i add the treatment+ medium to give final 200uL/ well. so i don't have to discard old media before treatment.
thank you
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