Cleaning sequencing gel plates
bornemd at aa.wl.com
Wed Nov 17 15:07:54 EST 1993
In article <2cdhvo$s4n at gazette.bcm.tmc.edu>, jamess at mbcr.bcm.tmc.edu (James
S. Sutcliffe) wrote:
> A convenient method for "siliconizing" sequencing plates is to use something
> called "RainX", a commercially available
[ ... ]
> chemicals, and doesn't have to be used in a hood. We routinely clan our
> plates with alconox detergent, rinse with water followed by 70% EtOH.
> Jim Sutcliffe
> Institute for Molecular Genetics
> Baylor College of Medicine
We used to do radioisotopic sequencing so we've used most of these methods
posted so far, but now that we use the ABI sequencer, it is very important
that our plates are optically clean (i.e., no absorbance/fluorescence at
the relevant wavelengths) in addition to the usual standard of cleanliness.
I think some of the lessons we've learned from cleaning plates and pouring
gels for the ABI are relevant and potentially useful to radioisotopic
sequencers so I'm sharing them here.
We scrub plates with a gloved hand and a sort of abrasive paste made with
Alconox and a very small amount of water (bad as this sounds the alconox
doesn't scratch the plates). The scrubbing is _VERY_ thorough and takes a
while. The plates are the rinsed very thoroughly (3-4 times) with
deionized water (we don't have distilled water), and the rinsed one final
time with 18 megaohm MilliQ water. The plates are propped up and allowed
to air dry. We then tape them up for pouring, put saran wrap over the top,
and store them like this to keep dust and lint from getting inside. No
ethanol, rainX, etc. since these can cause optical problems.
When the plates are this clean and dust/lint free there is nothing to
interrupt the smooth flow of acrylamide as you pour the gel, so it is
relatively easy to pour bubble-free 0.4 mm gels using a 60 ml plastic
syringe to dispense the acrylamide. When we disassemble the plates after a
run (using the handy-dandy Hoefer Wonder Wedge to prevent damage to the
plates) the gel remains intact and stuck to one plate so that it would be
easy to transfer to Whatman paper (this isn't necessary with the ABI). The
point of all this is that siliconizing or RainX'ing the plates is
unnecessary if one takes the time to do the cleaning job right.
We also mark the plates so that they are used in the same orientation each
time (top/bottom and inside/outside) to further protect the inside from
nicks and chips that can disrupt a smooth pour.
We've found it more economical to buy Alconox in single use packets than to
have half of it solidified in the bottom of their large box.
Cleaning the plates soon after use (before the acrylamide dries) makes the
job much easier.
The only downside of this method is that it's tough to get more than two
people to share the same standard of cleanliness so if each of your sets of
plates has to be shared more widely than this it might not be possible to
use this minimalist approach to cleaning and pouring.
Hope this helps,
Ann Arbor, MI
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