This was a busy week

So much stuff happened this week, that writing down the protocols for everything and explaining what they mean would take up enough space so I wouldn't have to do my blogs for the rest of the project (If I could write it all at one time). It started on Wednesday with day one of two of the western blot (a procedure to test if a specific protein was in the sample). This particular western blot was not part of my project, but since we would be doing one later, we figured it would be informative if I knew the process beforehand. It involved making samples with a lysate and water, putting it between heat blocks (exactly what they sound like) and then into wells in one of these:

along with ladders (not actual ladders), filled with buffer solution, then packed in ice an left to sit for 1.5 hours while running at 125V . After that was over, we had to make a "sandwich" of sponges, the gel that was in the contraption, filters and membranes. Also everything was soaked in buffer. Then the sandwich goes back into the western box which is again filled with buffer and packed in ice and left to sit for 3 hours on 125V. Then the membrane with the stuff on it is taken out and poured with Ponceau stain to check for protein. That's when I had to leave for the AMC Math competition, but the protocol for the rest of the day says that it then needs to have a 1 hour blocking with 5% milk (5g dry milk + 100mL TBST) followed by: Incubate in primary antibody (find dilution in AB datasheet, dilute in 2.5% milk) overnight in the fridge.

While that long three hour break was going on, we split the cells from our project. We basically took out the cell food, used Trypsin to break the cells from the wall of the flask, incubated it at 37 degrees Celcius for 5 minutes, then added cell food back into the flask, swished it around a bit, put it in a 50mL conical, broke up clumps of cells using a seralogical pipette, put in media into each of the new flasks, then 1mL of the cell-Trypsin-media went into the flasks.

And that was Wednesday.