0:01 [Music]
2:44 when placing the glass in the casting
2:46 frame make sure it's on a flat surface
2:48 so it'll seal tightly when placed in the
2:50 casting stand
2:53 [Music]
3:05 you may want to Mark the glass as a
3:07 guide so you know how much running gel
3:09 to add ideally Your Mark should be about
3:12 1 cm below the teeth of the comb you can
3:15 insert the comb to double
3:19 check once the APS and tet are added the
3:22 solution will start to polymerize so try
3:24 to add them as quickly as possible
3:26 [Music]
3:54 to add the running gel solution place
3:57 the tip of the PIP pet in the corner of
3:58 the space between the short plate and
4:00 the spacer plate it's a good idea to
4:02 leave a little bit of solution in the
4:04 pipet to avoid introducing bubbles into
4:07 the gel continue filling until the
4:09 solution reaches Your
4:12 [Music]
4:19 Mark adding isopropanol to the top of
4:21 the gel will keep the solution from
4:23 forming a meniscus within the glass
4:25 resulting in a very smooth surface don't
4:27 forget to remove the isopropanol layer
4:29 before adding the stacking gel
4:32 [Music]
4:38 [Music]
5:24 [Music]
5:34 [Music]
5:48 once the running gel solution has
5:50 polymerized and formed a plug you may
5:52 add the APS and Ted to the stacking gel
5:54 solution and then add it to the top of
5:56 the cassette typically you should allow
5:58 20 minutes for polymerizing
6:01 [Music]
6:17 [Music]
6:24 ation be careful not to damage the gel
6:26 while removing the isopropanol layer
6:31 [Music]
6:42 [Music]
6:50 remember that APS and Ted cause the
6:52 solution to polymerize so add them
6:54 immediately proceeding addition to the
6:56 cassette
6:57 [Music]
7:51 bring the level of the stacking gel
7:52 solution to the top of the short glass
7:56 [Music]
8:04 when inserting the comb some of the
8:06 stacking gel solution will spill over
8:08 wait until the solution is polymerized
8:10 before cleaning the acrylamide is
8:12 considered toxic until polymerization
8:16 [Music]
8:22 occurs when a gelatinous plug forms the
8:24 vile you are ready to remove the comb
8:27 from the
8:28 gel the wells can be a little tricky to
8:30 find Once the comb has been removed from
8:32 the finished gel so marking the first
8:34 well prior to removing the comb can save
8:37 you some
8:38 time be careful not to remove the comb
8:40 at an angle because this can damage the
8:42 wells
8:45 [Music]
9:26 you are now ready to run your gel
9:27 through electropheresis to examine your
9:29 sample good luck
9:33 [Music]
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