PAGE Races

Welcome to the PAGE races! I will give a quick introduction to all of our competitors before we begin our 1 hour journey through pores big and small. Ladies and gentleman please  fasten  your safety belts. We will be traveling at a comfortable 200 volts and do not anticipate losing cabin pressure, but in the event that we do....just kidding. Here are the samples and the lanes they were loaded in.
Lanes:
1. Brain (non-hematoma)
2. Brain (non-hematoma)
3. Brain (hematoma)
4. Brain (hematoma)
5. Blood (sheep heart artery/ new)
6. Blood (sheep heart artery/new)
7. Blood (sheep heart artery/old)
8. Blood (non-sheep)
9. Urine
10. Bio-Marker

And the winner is......I can't tell. The polyacrylamide gels that I am using are long enough for the tiny proteins that i am trying to extract. The protein that I am after is around 4,000 kDa. This gel only goes to 10,000 kDa. The good news is that the proteins are actually moving in the gel  which they were not doing with the previous SDS buffer I was using. This makes the smell of this new stuff worth it. The bad news is that the proteins are still a streak at the end of the plate. We have vertical electrophoresis tanks here in the lab which I can try. The only problem is that the gels cannot be purchased and polyacrylamide in it's polymer form, according to C3PO can "cause a very unpleasant death". I quickly imagined a bugs death and decided to research another way to run these vertical gels. I found a study that successfully used a 3% agarose gel (the bigger the percentage the smaller the pore size). My first attempt at pouring these gels was not successful. 1.8 grams of agarose is dissolved into 60 mL buffer solution. The solution is then injected into 2 panes of glass (about 8X11"). However, when I injected the gel it started to coagulate halfway down the glass. Oh, I almost forgot, the pieces of glass are so close together, the only thing I could find that would fit in-between them is an 18 gage needle. I also grabbed a used mini-gel cassette and super glued it together in order, and filled it with my new agarose solution to see if it can be reused. My second attempt at puring the vertical included the plates clamped with a rubber seal in-between the glass and placed in a water bath. This was also not successful due to water breaching the seal and compromising the gel. I have a new idea for attempt #3 but I will not ruin the surprise now :)
It was great to see you all in the lab today. Thank you Jeremy for all of your help this week (including all of your witty banter).
Also Thank you to Josh who has been so amazingly patient and accommodating. I appreciate the time you spend helping me pour through journal articles and combining all of the brainstorming sessions.