Hi, I need to know about cell counting for time point experiments-
I am working on Raw 264.7 cells which has 11 hours doubling time.
I use to seed 1 Million cells in six well plate which has capacity of ~1.2 million cells but they become 2 million before I done with infection.
I can control this by seeding the cells just 5-6 hrs before infection since Raw cells usually take 5-6 hrs to get attach.
But how to control time points?
I need to take out samples at 6,12,18 and 24 hrs.
I use to infect at MOI of 1:10, so I use to add same amount at one time only but for my 24 hrs exp. cells become more so I feel infection got diluted? How to control this?
One more thing, to handle this i also seed 1 million cells in 60 mm plate, but I am worried since cells are less and surface area is more, is it going to affect infection?
Why don't you try seeding cells at less density? 1 million cells in a 6 well plate makes this 100,000 cells/cm2 and in my opinion that is too much cell density for any cell line.
You can try lesser cell density i.e. 25000/cm2 (2,50,000 cells/well) as using this density you will get 5,00,000 cells by 11h and around 1 million cells by 24h.
I think this way you can control your time points.
I think that will also help with your MOI issue. You can try using a ratio for cells/cm2 of the flask.
Seeding cells/cm2 area of any flask or culture plate can resolve the issue of number of cells vs. infection.
- Badges
- Science topic
- Similar topics
- Physical Sciences
- Materials Science
- Plating