loading gels - what to use?

Bruce Roe BROE at aardvark.ucs.ou.edu
Sat Nov 2 12:27:04 EST 1996


Tamy writes: 
=> 
=> Lab ergonomics is a hot topic today.  
=> What alternatives are being used to load gels?
=> 
=> I have seen that ABI has recommended multichannel syringes with the XL 
=> upgrade to 48 wells.  I'd appreciate hearing about your experience 
=> using this system.
=> 
=> We are currently loading 36-well combs on the 377 and 373 with Gilson 
=> pipets and are finding wrist and fatigue problems...especially by the 
=> fourth gel of the day.
=> 
=> Many thanks
=> Tamy Hollingsworth
=> hollingsworth at immunex.com
=> 

Hi all,

	For about a year now we and others have been running 48 lanes on both
the 373 and 377's using a 48 lane comb file that became available in the public
domain.  This 48 lane comb file can be obtained by all through the web via URL:
	http://www.genome.ou.edu
under "Protocols".  You can also get there via ftp to:
	ftp.genome.ou.edu
I didn't do the hack to create this file so don't give me either credit or
blame.  I did hack the 24 lane file long ago to get to 36 lanes and was ready
to do the 36 to 48 lane hack, but someone else thankfully did it.

Down load the file if you want and put it in the ABI folder that's in the
system folder after un-binhexing it. However, you'll have to revert back to
Data Collection Version 2.0 rather than Data Collection Version 2.0.1 on
the 377 because ABI removed the ability to collect more than 36 lanes when
they upgraded the Data Collection software for the 377 from Version 2.0 to
2.0.1.  The 373 Data Collection software that works is version 1.2 as I recall.
Please, I can't distribute this software so please don't ask.  Sorry.

Yes, 48 lanes is a pain to load and also a pain because without the 388 data
point (aka XL upgrade) you have to spend the time to manually re-track the
data with the existing ABI software.  Hopefully the manually re-tracking will
not be necessary, I can always dream, with the XL upgrade.

Even without the 48 lane comb file you can run 48 lanes.  I wouldn't recommend
more than that because the 373 and 377 only collect 194 data points accross
the gel and that's 3 pixals/sample with one pixal between lanes.  It's truly
a pain but you can manually track the first 24, do the analysis and then go
back and manually track the second set of 24, to get the data from all 48
lanes.

Now, as for loading, with 48 lanes, the spacing of the combs corresponds to
the spacing in a 96 well microtiter plate offset by 3).  So until someone
(come on Lycor, make Elaine's autoloader and sell it for the ABI's, since they
don't seem to want to do it!!), we have three choices.

1. The Pipetman (Gilson or whoever) with the duck-bill flat tips.
2. A Hamilton syring (available from Fisher, VWR, etc) with the smallest
   needle or the tapered needle
3. A Hamilton 8 channel syringe (also available from Fisher, VWR, etc) with
   the smallest needles or the tapered needles.

The problem with the multichannel syringe is that the "datasheet" is difficult
to write because the samples from a microtiter plate are placed in wells that
are 3 wells appart.  Thus, you load 8 samples in wells 1,4,7,10,etc, (and then
another 8 beginning at well 25, and then the third set of 8 ending at well 46)
run on for 2 minutes, then load the 3 sets of 8 samples in wells 2,5,8,11,etc,
run on for another 2 minutes and then load the last 3 sets of 8 samples in
wells 3, 6, 9, 12, etc.  However, the time to load 4 gels with the 8 channel
syringe is less than a half hour, including run-on times.

We, and I'm sure others, are working on writing a program that would create
the "samplesheet" so you don't have to enter the data manually there.  However,
because each lab has different naming conventions this program would be
difficult to transport to other labs, which is why ABI probably hasn't written
one.  Once we get ours completed and tested, I'll put it on my ftp/web site
but you'll have to take it as is, warts and all, if you want it.  We should
have this done before the end of this month so visit my web site for it then.

Wishing you all well and happy sequencing.
Cheers.........Bruce
***************************************************************************
Bruce A. Roe, Ph.D    Department of Chemistry and Biochemistry
                      University of Oklahoma, Norman, OK 73019-0370, U.S.A.
Phone: (405) 325-4912 or 7610;  FAX: (405) 325-7762;  e-mail:  broe at ou.edu
********************** http://www.genome.ou.edu/ **************************




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