Hey All,
I just wanted to check if anyone has observed differences (vigor and viability) in thawed THP-1 cells at varying initial freezing concentrations (Cell/ml). In general I freeze down stocks at ~10 x10^6 cells/ml for other cells. Would this be an appropriate concentration for THP-1s? We've had some issues with this cell line in the past.
Thanks!
Dear Nathan,
I did not tested several cells concentration , but I tested differences in the temperature of freezing for THP 1 cells: they have better vigor and viability in -80°C than in Nitrogen storage. I hope it helps you.
Best regards
Cristián
Number of cells are fine. It's your technique which matters. This is the way I store the cells and put them back in culture, and never got problem
1. Centrifuge cells
2. Pour off medium completely
3. Add 900 ul FCS to the falcon tube and gently pipette up and down to resuspend cells
4. Transfer cells and FCS to cryotube vial with 100 ul DMSO
5. Mix with mixer and immediately put it in cryobox or wrap in tissue, and transfer to -80 C freezer
6. Leave overnight
7. Transfer to liquid nitrogen box for storage
Also you need to be very quick when putting them back in culture:
1. Put RPMI in 50 ml falcon tube
2. Retrieve cells from liquid nitrogen
3. Quickly add some RPMI and pipette out the suspended cells into the 50 ml falcon tube, and centrifuge at 1200 for 5 min
Typically I freeze 4x10^6 THP1 cells per ml in 90% FBS + 10% DMSO. I have not tried multiple cell concentrations, but a word of advice is that these cells typically take longer to recover from cryostorage than other cell lines I've worked with. Using 20% FBS in the culture media for 2 days following thawing cells helps them recover quicker
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