I use the CBS Scientific gel assembly apparatus (http://www.cbsscientific.com/verticalmini-gelelectrophoresissystemaccessories.aspx) to make polyacrylamide gels. Lately, I have been experiencing a lot of wells collapsing such that the walls fall over and it is nearly impossible to get them upright again.
The slanted, fallen over well walls cause sample to be loaded at an angle which results in samples overlapping in the resolving gel. This issue has made interpreting gels very hard as individual lanes become nearly impossible to discern.
Does anyone know how to prevent the well walls from collapsing? Any technical advice would be helpful and appreciated.
I think the main problem with removing the comb is that you form a vacuum in the wells when you pull the comb out and this is often what distorts the wells. So I found that I can get rid of the vacuum by pushing the top/holder of the comb a TINY bit forward (away from the back plate) before I remove it. You have to be careful to push far enough to allow a small amount of air into the wells but not so far as to allow air between the plates and the gel. The tiny bit of air that rushes into the wells allows me to remove the comb without a problem. I also find that wetting the assembled gel (plates, comb, the whole thing) under a tap just before I remove the comb seems to help (it might be my imagination but I think older/drier gels are more problematic than fresher/moist gels).
yaa....it is a situation based problem.
when your gel is properly made then remove comb just before loading, avoid vacuum formation, u can increase inter-well space and always handle very gently.
well collapsing may also uccur due to less % of acrylamide or misfunctioning (low quality) of chemiocals used.
You can store the gels 1 day at fridge, with glass plates and combs wrapped in cling film. When you use them, you must put the gels in the electrophoresis tank, with running buffer. After that you can remove the combs without any problem.
If the gel had properly polymerized, I usually find benefits in pulling out the comb slowly, while keeping the gel in an inverted position. If collapsing still occour (but usually do not), use a long tip to enter between the glasses and try to restore it in the proper position.
I always pull out the comb just after the electrophoresis chamber is put together and filled with running buffer (fully between the two gels). That way the buffer will prevent the collapse of the wells. However, if the gel is not polymerized properly the wells will collapse anyway.
i had this problem recently. and i did these tips:
first, i stirred the mixture just for 1 to 2 minutes after adding TEMED.
then, i poured it between glasses and put the comb in.
after that, when the gel was polymerized, i filled the inner tank with running buffer and pulled the comb out just before injecting the samples into the wells.
it works for me.you can try it.
good luck
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