I'm working on SH-SY5Y viability test with MTT assay, using vitamin C and H2O2.
In our lab protocol, after 30-48h from cell seeding it reaches 70% confluency and gets ready for cell deprivation. Also, I learned that deprivation must start when cells are at 70% confluency.
However, I found out that many research studies have incubated sh-sy5y for 24h and treated the cell right away.
I've seeded sh-sy5y more than the research's density, but they didn't reach 70% even after 30h.
I'm wondering since sh-sy5y has a long doubling time (almost 3 days), is it possible for sh-sy5y to reach 70% confluency after ~48h from seeding?
Then should I wait more than 48h after seeding?
t sounds like you've encountered a bit of a conundrum! The doubling time of SH-SY5Y cells does indeed play a role in the confluency timeline. Given their slower growth rate, waiting for around 48 hours might not be sufficient for reaching 70% confluency.
It's a good idea to adjust your protocol based on the specific characteristics of the cells you're working with. If 48 hours hasn't brought them to the desired confluency, you might want to consider extending the incubation time.
You could try waiting for a bit longer, perhaps up to 72 hours or even more, to see if the cells reach the desired confluency. Regularly check their growth progress and adjust the timeline accordingly.
In research, it's not uncommon for protocols to be adapted based on the specific behavior of the cells being used. If you find that a longer incubation time is necessary for your SH-SY5Y cells to reach 70% confluency, it's perfectly reasonable to make that adjustment.
Remember to monitor the cells closely, and when they reach the desired confluency, you can proceed with the cell deprivation and treatment as outlined in your experimental design.
A.B Bayazid Thank you for your previous responses :)
Despite seeding a substantial amount of cells to achieve 70% confluency within 24 to 48 hours, considering the doubling time is around 3 days, is it actually challenging to characterize the cell growth curve as being in the exponential phase?
There are many contributing factors that can influence the doubling time of SH-SY5Y. It's documented that use EMEM full medium for SH-SY5Y cells.
If your cell stock is years old then make your cells grow slowly, for one or two weeks and then after a few passages, they will start to grow rapidly.
Same thing I experienced myself while working with SH-SY5Y cells.
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