Hi, In our lab we store P. aeruginosa isolates in Tripticase soy broth with 15% glycerol at -80oC, but almost when we culture those stored isolates they do not growth or only are a few colonies. So do you have a good method to store this bacteria?. Thank you
Hi,
In our lab we grow the Pseudomonas aeruginosa strain in TSB to log-phase (OD580 of 0.300) and then add glycerol to a final concentration of 20% to freeze at -80°C. With this conditions we have an excellent recovery. Maybe your problem is the very low glycerol concentration you're using.
Hi Paul. Try storing them on cryobeads. This is what I did with my Ps. aeruginosa strains and I never had a problem.
Hello Paul,
Try to maintain P. aeruginosa into LB broth in cryogenic vial with 50% glycerol at −80°C.
Hi Paul,
We stored the p. aeruginosa isolates in 30% glycerol in triplicta. You should however replete them regularly (every two months or so.
P. aeruginosa can be preserved in frozen stocks of either 20% glycerol or 10% skim milk that can be stored at −80°C
Hi, in my lab we preserved P. aeruginosa in Special Gelose vials at 4°C for 2-3 years (Blood agar base 20 g, Bacteriological agar 15 g, Beef extract powder 1.5, dH2O 1,000 mL, pH - 7.2)
Yes, if you increase glycerol % to 20 , there will be a difference
The rate of bacterial recovery depends on: 1. Initial concentration of bacterial cells; 2. The presence of protective substances in the culture media; 3. The depth of freezing and 4. The method of recovery. So, I think you would better to precipitate the bacterial cells (4000 g for 10 min) and discard the supernatant. Afterwards, add 2 ml freshly prepared Trypticase Soy Broth (TSB) containing 30% Glycerol and store them in liquid nitrogen. For recovery, it is better to use TSB containing 0.5% yeast extract for a period of time longer than normal (2-3 days at opt temperature).
Lyophilization (freeze drying) could also be helpful.
I freeze them in a maximum medium (ex. PCA), 20% glycerol and after 10 years they are still viable.
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