Dear community,
I need to obtain viable cells of my bacterial isolates during their exponential phase. Exponential phase is known from growth curve using spectrophotometer. Thus I'd need to centrifuge the bacteria in medium if I want to obtain only the cells. I am wondering what is the maximum speed and duration of centrifugation to obtain viable cells? My bacterial isolates are still unknown but they are of cocci-shaped and mostly Gram-positive bacteria.
Best regards,
Nur Shabrina.
Well you can try same speed as is used for cell centrifugation: 1200 rpm, 10 minutes at 4C. You can make mistake while trying to optimize your conditions.
If they are Gram-positives, they should survive basically almost all g-forces you can realistically put them through. Everything below 20,000g should be no problem at all, even for normal Gram-negatives. So 2-3 min at 20,000g should get most of them in the pellet, depending a little on your media.
I would not cool them though, as some bugs do not like severe temperature changes. Once you have isolated them, you can then run some tests.
I would also feel a high g force will be fine (upto 15000) without much harm for gram positive cocci, however, I believe a temperature between 4-8 deg C may yield more viable cells
Dear Dr.,
As you have mentioned that the bacteria are in their log/exponential growth phase, centrifuging these at high rpm may damage the growing cell wall and membrane due to higher shear and stress exerted by higher centrifugal force. So it is suggested that you use 1500-2000rpm for 6-10 minute at 8-10 degree celsius (the lower temperature may inactivate your actively growing cells). This temperature will definitely ensure greater viability (instead of working at 4degree).
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