Cleaning Electroporation Cuvettes (summary)

David Nunn David_Nunn at qms1.life.uiuc.edu
Wed Mar 17 16:16:09 EST 1993


Sorry I didn't post this summary earlier but here it is.

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Hello!
We have been using the BioRad Cuvettes for bacterial
electroporation and have very good results using teh 
following method.
1) Immediately after using the cuvettes, we rinse them
in bleach followed by lots of distilled water. (This is 
to kill the left over bacteria prior to disposal down
the drain)  After the water rinse, we rinse the cuvettes
in 95 % ethanol and let them dry.  We store them dry until
we have enough to process.

2) Fill the cuvettes with 0.25 M HCl and let stand at room
temperature for 15 min to 2 hr. (This step depurinates any
DNA left in the cuvettes)

3) Rinse the cuvettes in distilled water and place them in  
a boiling water bath (fresh, clean, distilled water) for 
about 10 min.

4) Remove them from the bath and immediately rinse them in
95 % ethanol and let them dry upside down. (We have found this
to be sufficiently sterile for bacterial work).  The caps are 
rinsed in 70% ethanol and dried in a sterile hood but this is 
probably not neccessary for just bacterial work.

One of the most important things to extend the life of the 
cuvettes is to not store them for any length of time in the
presence of moisture.  (the aluminum electrodes corrode rapidly)
Also, the boiling water bath is essential to hydrolyse the 
depurinated DNA. A postdoc in our lab ommitted that step and 
had problems with unwanted colonies in subsequent electoporations.
Using the above method, we have been able to use our cuvettes
at least 5-10 times before discarding them.

Good luck!   
X-News: crcvms bionet.molbio.methds-reagnts:3154
-----------------------------------------------------------

We 
-rinse them a few times with dH2O
-fill them with 0.1N NaOH and let them sit for about 5 minutes. Note the
scrubbing bubbles!
-Rinse well with dH2O
-Rinse two or three times with 95% EtOH.
Finally, dump out most of the EtOH, replace the caps, turn them upside down
and lean them against a support to dry.

This is quick and effective. We usually keep squirt bottles full of these
solutions near the sink. It's also possible to hold as many as eight
cuvettes
at once during this protocol by lining them all up and holding the bunch
between fingers placed like bookends at the end of the line.


Christian E. Fritze                   |                           
AOL:geneman
University of Chicago                 |              
fri0 at midway.uchicago.edu
Molecular Genetics and Cell Biology   |   "No one ever died of laughing"
-M.B.
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I've not had to recycle my EP cuvettes yet, but I have several hundred of
them
lying around in case times get tough!  I noted in a product review in
Nature
347, page 310 a year or two back that an ABI robotic PCR machine uses a
dilute
nitric acid wash to remove DNA from its single PCR plate (I forget whether
it
was aluminium or not, but I suspect so).  I reckon that if nitric acid can
clean a plate sufficiently that there is no carry-over of DNA between the
PCR
reactions then it ought to be effective at cleansing EP cuvettes.  Perhaps
it
is worth a go - let me know if it works!

Cheers,

Martin Kennedy

-----------------------------------------------------------



	We place our used cuvettes into a Branson sonicator and add a mild
detergent (joy or other dishwashing detergent).  Turn on the heat to 50C
and
sonicate for 45 minutes.  The cuvettes are then thoroughly washed with dH2O
and finally placed on a cuvette washer and washed with 70% EtOH.  We dry
them
in a vacuum oven with no heat and store them in a sealed container.  We
have
not found any evidence of DNA or bacterial cell contamination by this
method.
One note, the cuvettes do not last indefinitely, and in our hands are good
for
about 6-10 electroporation cycles.  After this, they usually have so many
cracks as to make them unsafe.  Hope this helps,

		"Ignorance is a
		 voluntary misfortune."
		  -- Nicholas Ling
		
		Bob (Belas at mbimail.umd.edu)
-----------------------------------------------------------

We just soak the cuvettes in 50% ethanol/water overnight and then expose
them
to UV light overnight the next night (we have a culture hood).  This seems
to
work fine as we haven't had any problems so far.

Any other ideas?

Dubear

-----------------------------------------------------------

I am very interested in this subject!  I presently am washing my cuvettes
2X in water and then soaking them overnight in 4N HCL.  I then rinse them 
under de ionized water for 15 minutes to 1 hour.  I then sterilize and use
normally.  I hae repeated this procedure a number of times with no loss
of transformation efficiency.  The acid breaks the DNA backbone into tiny
pieces and thus destroys it.
	If you get any other suggestions please forward them to me!
Thanks
Kirk Schnorr

-----------------------------------------------------------
Wash X6 with dH2O
Zap again
Wash X6 with dH20
Wash with EtOH
Store dry.

No carry over of DNA, no degradation of cuvettes.  An excellent method.

Anthony Davies
Institute of Virology
Oxford UK


-----------------------------------------------------------
David Nunn, Ph.D.
Department of Microbiology
University of Illinois
Urbana, IL 61801
(217) 333-6131



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