It is great to be back in the lab. This semester I will
continue my exploration of the Beta-Amyloid proteins and antibodies associated
with Alzheimer's Disease. I was hoping to be able to obtain these proteins
and/or antibodies but they are extremely expensive. The protocol that I used
last semester extracted proteins from fish, but did not extract proteins from
the brains or blood. So my first step was finding a protocol for protein
extraction from brain tissue. Once again, the solutions needed for the extraction
protocol that appeared to yield the highest results are very expensive. I have
combined a few different protocols and with the help of Matt and Josh I am
ready to begin this journey. My first challenge came when my protocol called
for a pH 6.8 Tris HCl solution and the Trisma base we have in the lab has
a pH of around 9.8. With the help of a pH meter (which needed to be calibrated)
and some HCl I achieved my first mission. Next I needed to make an SDS buffer
which was simple to mix but one of the smelliest things I have ever been
exposed to. Beta- mercaptoethanol is the worst smelling compound that I have
ever smelled (the rest of the lab will agree, love you guys). The next step of this protocol was to pulverize the brain tissue in a ball mill grinder (which, again is very expensive). After a brief meeting with our local lab droid (C3PO) I decided to simulate a ball mill grinder. This meant placing 81 ball bearings (from the bearings on my roller skates) into a falcon tube along with the brain tissue and 250 uL of SDS buffer. The falcon tube was placed on the vortex for 8 minutes. Next week I will be preparing samples, loading PAGE gels and performing electrophoresis to see if this smelly protocol works.
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