plate life

Marty Sorry at NotTo.Day
Tue Dec 10 17:29:58 EST 2002

In article <WqOH9.63$P4.9643 at news.uchicago.edu>,
 "EK" <khatipovNO-SPAM at NO-SPAMuchicago.edu> wrote:

> There is a wonderful methods book Methods in General and Molecular
> Bacteriology edited by Philipp Gerhardt. Here is the link to amazon.com with
> the reference
> http://www.amazon.com/exec/obidos/ASIN/1555810489/qid=1039118552/sr=2-1/ref=
> sr_2_1/002-1939933-5691258
> You will find several methods  of preservation of microorganisms. The best
> is to use the method recommended for your particular species.
> There are other books you can look for in the library, too.
> If you are working with sporulating fungi, I guess you could store spores
> for pretty long at -20 or -80. No freeze drying would be required. However,
> again, you should consult established protocols first.
> Another source of useful information would be websites of type culture
> collections like www.atcc.org or www.dsmz.de
>     Emir
> "Sara Caldwell" <jnskc at uas.alaska.edu> wrote in message
> news:3DEFA088.38E384F3 at uas.alaska.edu...
> > Everyone,
> > Thank you for all the great information!  Thankfully, we already had the
> > plates in plastic bags.  I expect when we have to make new media we will
> > switch to agar slants and wrap the caps in an extra layer of parafilm.
> > I just hate wasting materials.  If the plates do last for even two
> > months then I won't have to toss them out unused. :)
> > Your replies actually generated new questions.  For instance, a freeze
> > dryer was uncovered downstairs.  What would be the protocol for using
> > one to freeze bacteria?
> >
> > Sara Caldwell
> > University of Alaska Southeast
> >
> > P.S.  Are people finding that spammers snag addresses off this board?
> >
> > JEDilworth wrote:
> > >
> > > Not until Spring, 2004. When we made home-made media without blood
> > > enrichment in our medical microbiology laboratory we used to put an
> > > outdate of six months on them. You can try subbing organisms on to them
> > > periodically to see if they still work. Even if you wrap them in
> > > plastic, the plastic will breathe and the plates will end up getting
> > > dried out.
> > >
> > > How are you keeping the organisms alive? Are you planning on using this
> > > media? It will work for awhile, but you'll seriously need to consider
> > > freezing at -70 Degrees F (or minimally at -20 degrees F but that
> > > doesn't work that well), or subbing weekly to media. At about the six
> > > month mark you'll probably be making a new batch of media to sub onto.
> > > Why not make some slants? Those will store your subs better. What
> > > organisms are you saving?
> > >
> > > I'm sure other people on the group will have ideas also.
> > >
> > > Judy Dilworth, M.T. (ASCP)
> > > Microbiology
> > >
> > > Sara Caldwell wrote:
> > > >
> > > > We poured too many plates for microbiology this term.  We want to keep
> > > > the cultures we have alive till spring of 2004 when the class is next
> > > > taught.  How long will the following refrigerated un-inoculated plates
> > > > keep?   Brain Heart Infusion, MacConkey, Nutrient, and Tripticase Soy
> > > > Agar plates?


   For storage of most general laboratory strains of bacteria, I'd 
recommend you add about 0.5 ml of healthy log phase growth to 0.5 ml of 
2X TSB w/30% sucrose. Then put the cryovial in a -80 C freezer. This 
will work pretty well for storage for several years. You'd be even 
better off if you have access to liquid nitrogen storage (gas 
phase...not liquid phase). The gas phase (above the liquid level) is 
usually about -145 to -175 C and will preserve the cells for many many 
years. This may not work for some of the exotics, such as halophiles or 
environmental anaerobes, but usual teaching and qc strains will do 
pretty well. 
   Weekly subculturing is not good for long-term stability. Genetic 
drift and selective action by the medium may occur. 
   We lyophilize many bacteria using serum vials with rubber stoppers 
and aluminum crimp seals. These are dated for 5 years. We also 
lyophilize bacteria and seal them under vacuum in 100% glass. These have 
no expiration dating...they last a long time!
   I would advise you to stay away from freeze-drying unless you really 
are anxious to spend a lot of time learning about the process. You'd 
have to check out the refrigeration system to make sure it could acheive 
-60 to -80 C for the condenser, and the vacuum pump would have to be in 
good condition also. If the equipment is in good shape, the actual 
process isn't very difficult depending on the capability of the machine 
(manual or automatic, etc...).

Good Luck,

Martin Gross
Technical Manager
Media Prep and Cryopreservation
American Type Culture Collection
mgross at atcc.org

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