The medium I use is: IMDM, 10%FCS, P/S, 7.5% Sodium Bicarbonate, 0.1mM b-mercaptoethanol
Cells double their number in 24h after thawing, but then hey do not expand more (actually the number decrease). However they look healthy and after some days start to get attached.
Thanks
Did you change medium after plating? Or remove the freezing additive by changing medium by pelleting the cells and resuspend in fresh one?
Hi Jordi,
Is 7.5% sodium bicarbonate not very high? I mostly use it for mAb production in 5L.
On the other hand could you please give more information about your thawing method, as Attila mentioned?
Best
1.Did you ever used this media ?
2. Did you prepare media with yourself or used commercial media ?
3. What type of mouse myeloma cells for fused ?
Think you must add extra glutamine to the medium. Hybridomas particularly sensitive if glutamine concentrations are low. You can buy this from most media suppliers eg. Glutamax etc. that comes as a 100x concentrated solution.
Jordi,
Have you tried any other media (RPMI, F/12, DMEM, etc), with or without with that percentage of sodium bicarbonate, with the same cells?
Best
Jordi!!! I asked here at irsi to the people working with hybrydomas
They use more sera (20%) unless you have spend time adapting the hybridoma to grow with less sera
Also, if the hybridomas were frozen in HAT medium (were you select the hybridomas)
you should use media with more sera (20%) and Hypoxanthine+Thymidine the H and the T of HAT
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