Reused Electroporation Cuvettes?
George W Chacko
We have a BTX electroporator and I wondert if anyone has tried
re-using the cuvettes for it? I'd be most grateful to hear of your
experiences.
Thanks
GC
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: gch...@magnus.acs.ohio-state.edu (George W Chacko) writes:
: >We have a BTX electroporator and I wondert if anyone has tried
: >re-using the cuvettes for it? I'd be most grateful to hear of your
: >experiences.
: They blow up. This is very exciting. Kind of ruins your experiment.
: Actually, this is true, but also, I have reused cuvettes. As soon as you
: finish using them, wash them thoroughly with clean water, and let them
: try. Probably best not to use ethanol on them. They really do expload
: (actually, just arc an a rather noisy way and pop the cap off; this is so
: startling that it is very difficult for me to electroporate now without
: covering my ears and closing my eyes, but I've always been leary of
: blowing up baloons.) A good reason not to reuse cuvettes is that you
: will contaminate your next transfection with plasmid from your previous
: transfection; at low level but it will likely be there.
: --
: Richard Vogt (Internet: vo...@biol.scarolina.edu) (803) 777-8998
: Department of Biology, University of South Carolina, Columbia SC 29208
I don't use the BTX electroporator, but reuse the BioRad cuvettes by
first rinsing with water, then 1 hr in 30% HNO3 to hydrolyse any residual
DNA, wash again with water, then fill with 95% EtOH overnight, empty and
dry under vacuum.
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Dick Vogt
>We have a BTX electroporator and I wondert if anyone has tried
>re-using the cuvettes for it? I'd be most grateful to hear of your
>experiences.
They blow up. This is very exciting. Kind of ruins your experiment.
Nancy Lemieux
Yes, I have re-used electroporation cuvettes. The first time I tried this,
I just washed with ethanol, then water, and let them air dry ( I must point
out that I didn't use the cuvettes again until the following month or so).
When I re-electroporated with very low quantities of plasmid, 2 out of 3
transformants had the plasmids from the previous transformation. However,
most scientists learn from their mistakes, and I tried a new method where I
washed several times with water, and then rinsed the cuvettes with ethanol
and let them dry. My negative control on that one (E. coli without any DNA
added) gave absolutely no colonies, so I'm not really worried about low
levels of contamination. Of course, if you're really paranoid, you could
always try treating the cuvettes with UV light, or H2O2, or some such
treatment which destroys DNA.
Good luck,
Nancy Lemieux
Dept. of Biochemistry
McGill University
Montreal, Canada
n...@binkley.cs.mcgill.ca
ell...@nickel.ucs.indiana.edu
have been reusing both with no problem about 10-12 times each. VWR sells
the BTX cuvettes and were the next ones I was going to use due to discounts
etc.
The way we reuse them is as follows:
Rinse immediately with water, the 10% bleach for 10 minutes, rinse with
water thoroughly then 75% ethanol three times, put foil on top and put into
a 70 degree oven overnight, cool and store at 4 degrees so that they are
always prechilled and ready to use.
Our lab has had good results and no cross contamination between samples.
We also did a comparison between brand new cuvettes and reused ones to
make certain that there would not be a decrease in transformation efficiency
and the results from this showed that the transformations produced nearly
the same number of colonies plus or minus 5 colonies with the same tube
of cells.
Any cuvettes that I have been mildly suspect of, I put 50 ul of 10% glycerol
and do a "mock" electroporation to see if the cuvette arcs. Basically you
just need to look for the insulating plastic to have cracks in it between the
two metal sides, or look at the bottom of the cuvette which usually is very
cracked up looking. These two characteristics in 11 of 12 cuvettes meant
that the cuvette would arc if used. After a while you can spot the bad ones
without having to test them. It really saves on dollars and isn't that
cumbersome to do. Be careful not to let the bleach sit too long or it will
corrode the metal--only use 10% bleach---10 ml of standard household bleach
(5.25% sodium hypochlorite) plus 90 ml of water.
Hope this helps.
Ellen M. Quardokus
Indiana University-Bloomingto
Jon Visick
>We have a BTX electroporator and I wondert if anyone has tried
>re-using the cuvettes for it?
I have re-used cuvettes for the Bio-Rad electroporator virtually
indefinitely
(at least 10-20 times each) with no trouble; however, _very_ thorough
rinsing
is an absolute essential. My method: soak 20 min. or so in 10% bleach
(I use
them for bacteria exclusively, so want to be sure to kill all remaining
cells),
then rinse about 5x with water, 2x with ethanol and then 10x (no kidding,
ten
times) with the cleanest water you have available. They have never
"popped"
on me since I started this procedure--except for when I screw up the
actual
buffers/conditions! :)
eric c. anderson
<vis...@ewald.mbi.edu> wrote:
>
> >We have a BTX electroporator and I wondert if anyone has tried
> >re-using the cuvettes for it?
>
the protocol which our lab has developed for the Bio-Rad cuvettes is as
follows:
- immediately after use, wash for 5 min. with Milli-Q water.
- rinse 1X (and _very_ briefly) with 50% bleach solution.
- rinse 5-10 min. with Milli-Q water.
- UV irradiate for at least 5 min. with cuvettes placed upside down on the
light box.
i did not perform the controls for this protocol, a now gone grad student
did, but my experience has been that transformation efficiency is not
significantly different than with new cuvettes, and contamination has not
yet been a problem.
eric
--
BIG AIR...
NOT BIG PANTS!!!
---------------------------------
eric c. anderson
ande...@pharmdec.wustl.edu
660 s. euclid box 8103
washington univ. school of medicine
dept of molecular biol. and pharmacology
st. loser, misery 63110