Summary: Growth inhib. from detergent

David Kling dkling at warren.med.harvard.edu
Wed Dec 29 16:48:19 EST 1993


Dear netters:

Here is a summary to the query:

Has anybody observed growth inhibition in E. coli due to detergents used
for washing glass-ware?



David,

There is a detergent called 7X which I believe is specifically used
for washing glassware to be used in cell culture. As the detergent
itself appears to be relatively non-toxic, extensive washing to remove
every last bit is not as critical as with other detergents.

Dean Lee
Dept of Micro
LLU
dLee at ccmail.llu.edu


We have found that trace quantities of detergents can inhibit growth. The
effect
is especially noticable when making competent cells. We don't acid wash,
washing
a few times in ddH20 seems to be effective


Sam Aparicio
saparici at med.cam.ac.uk

Hi David,

The only thing I know that may be of some help is that the old Hanahan
procedure for making competent cells warns about traces of detergents in
glassware lowering competency.  One step in that protocol is to autoclave
the flasks with water, and then throw the water out.  Whether this really
helps or would help you I don't know.  If I had to guess, I think a
non-ionic detergent like Triton might be less harmful than the usual
Alconox.
Good luck.

Phil Carl

Sounds simple, but how is glassware being washed?

'tsi best for ALL glassware to be rinsed IMMEDIATELY after use, couple
times with warm -> hot tap water. This way junk is more likely to actually
GET WASHED OFF!

Also, if liquid detergent (such as for automatic washers) ... beware, some
are hard to rinse properly, and safest thing is to use VERY HOT WATER
rinse cycles (3-4 rinses of at least 2 minutes each) after wash cycles,
followed by dH2O rinses. 

If powder used, same precautions needed ... HOT water rinses AFTER wash
cycles followed by dH2O rinses.

Same care neede if washed by hand (wear gloves due to HOT water needed),
can rinse 2x in hot tap water then soak TOTALLY submerged in dilute
detergent sol'n in sink/tub ... many days can go by ... key is to NOT
allow surfaces to DRY out, thus total submersion needed with air bubbles
removed ... wash with soft brush ... rinse well with HOT TAP water then
few times with dH2O ... 

use of HOT water rinses (even with "hard" tap water), also helps reduce
spotting if that is ever of concern.

As result, we've been able to successfully use ANY detergent (we've used
at least dozen different brands, types, etc over past few years ... in
all, we've found rinsing steps to be most vital ... BEFORE washing
(immediately after use, especially for agar, agarose, bacterial cultures)
... and after washing ... to remove any residual detergents.


... I wash many of my own glassware (when time permits), otherwise we have
a dishwasher who is always striving for perfection ... thus consults with
us regularly (we've always considered her part of our research lab as
result ... a bit of a keener I might add) ... thus, we've always managed
to solve any problems that appear to stem from glassware cleaning (and
management of students and other lab workers in order to be disciplined
enough to follow "simple golden rule of rinse after use, otherwise that
piece of glassware may never get quite clean enough again).

cheers,
                            ********************
Andre Hamel                              email: hamel at ccu.umanitoba.ca
Manitoba Veterinary Services          lab tel.: (204) 945-7630
545 University Crescent,                   FAX:(204) 945-8062 
Winnipeg, Manitoba, 
CANADA   R3T 5S6            ********************


Hi David,

>I think the detergent we are using in the lab to clean our side-arm flasks
>is inhibiting the growth of bacteria, most notably, E. coli strain
>DH5-alpha.  Has anybody encountered this phenomenon?  Are there detergents
>available that do not inhibit bacterial growth or must we either acid wash
>our flasks or not wash our flasks with detergents?  Please respond directly
>to my E. mail address,I will post the list of responses together.

I regard detergent as one of the nastiest contaminants in the lab!  For all
glassware, except those that have contained phenol or other difficult to
remove
substances, I simply recycle by rinsing in water.  
Cheers,

Martin

NNNN   NN  Martin A Kennedy (E-mail = mkennedy at chmeds.ac.nz)  ZZZZZZZ  
NN NN  NN       Cytogenetic and Molecular Oncology Unit          ZZZ
NN  NN NN           Christchurch School of Medicine            ZZZ
NN   NNNN              Christchurch, New Zealand              ZZZZZZZ
       Phone (64-3)364-0880  Fax (64-3)364-0750

Subject: growth inhibition from detergent?



Dear David

We came to a similar conclusion in my previous laboratory, where we worked
on
ATPase mutant strains of E. coli - Doing growth yields on limiting glucose
or
succinate, or growth rate curves.  Our answer was to avoid detergents
entirely,
and to be fanatical about rinsing the flasks with distilled water
immediately
after use.  I suspect the occasional acid-washing would be beneficial,
although
we did not bother.  Hope your problems disappear soon.
Cheers
Rob.S.

          _______________________________
          |      Robert G Solomon       |
          |  c/- Dept. of Biochemistry  |
          |   University of Cambridge   |
          |      Tennis Court Road      |
          |      Cambridge CB2 1QW      |
          |     (0223) 333662 phone     |
          |     (0223) 333345 fax       |
          _______________________________

"When the only tool you have is a hammer, every problem looks like a nail."


hi,
i have not noticed this phenomena myself, but i have noticed that dh5-alpha
are
often very slow to grow, particularly with large (or otherwise) inserts.
otherwise, i cant think of anything.  i guess you do wash the flasks
afterwards
well?

best of luck

--
Rod Bonfiglioli, Waite Agricultural Research Institute,
University of Adelaide, South Australia.
e-mail rbonfigl at waite.adelaide.edu.au

 I've regularly grown E. coli DH5alpha in flasks that were washed
with the usual powdered dish detergents or with Sparkleen (Fischer)
without any problems, as long as the glassware was extensively rinsed
after washing.  I generally found that after being washed in a regular
dishwasher the glassware needed to be run through a second rinse cycle AND
hand-rinsed in distilled water to get rid of all the soap, and that was
with using about 1/3 of the reccomended amount of detergent.



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