I am looking into the plates that are optimal to re-stimulate murine splenocytes in, and subsequently use the same plate to perform ICS in. There are non-tissue culture treated plates available and tissue culture treated plates, and it seems that the TCTP are for cells that adhere, while the non-TCTP plates are used for cells that stay in suspension.
Now I am reading around and there are some people working with mouse splenocytes that are adherent and some people work with suspension cells. I know my cells are usually in suspension, but some will always adhere to the plate surface.
I would like to know what other people use in the field, so I can make a better decision on which plates I should use. Thanks :)
The non-tissues culture treated plates are primarily used for in vitro assays, such as ELISA, estimation of protein concentration, serial dilution of substances and not meant for tissue culture.
Cells - especially primary cells such as splenocytes - can be very sensitive to the tissue culture dishes. Tissue culture pre-treatment provides optimal conditions not only for the adherent cells but also for cells in suspension, such as splenocytes.
The non-tissues culture treated plastic often has stimulatory potential on its own and then you will not really know if your splenocytes responded to the tested substance/antigen or if the response was due to a plastic. Also, the cytokine expression pattern can be influenced....
You can of course use your time and Money of your laboratory to do comparative experiments and to define precisely, which plastic is "safe" and which not, but I guess that it would be a project on ist own.
Good luck with your research!
The non-tissues culture treated plates are primarily used for in vitro assays, such as ELISA, estimation of protein concentration, serial dilution of substances and not meant for tissue culture.
Cells - especially primary cells such as splenocytes - can be very sensitive to the tissue culture dishes. Tissue culture pre-treatment provides optimal conditions not only for the adherent cells but also for cells in suspension, such as splenocytes.
The non-tissues culture treated plastic often has stimulatory potential on its own and then you will not really know if your splenocytes responded to the tested substance/antigen or if the response was due to a plastic. Also, the cytokine expression pattern can be influenced....
You can of course use your time and Money of your laboratory to do comparative experiments and to define precisely, which plastic is "safe" and which not, but I guess that it would be a project on ist own.
Good luck with your research!
Thanks Agnieszka! I guess I will have to try both type of plates, and see if my results are different :)
Leroy, when doing ICS on mouse splenocytes, we place the cells directly into FACS tubes, add antigen and transport inhibitor, and culture for 6 hours. We then perform the staining right in the tubes.
For high-throughput analysis, we use non-treated, round-bottom 96-well plates for the stimulation and staining. After completing all of the staining steps, we then transfer to FACS tubes to run in the cytometer.
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