Matt Boyle, Entry #5, Horse Round 2 and Last Day

After some thorough research, I found that equine IL-4 is nearly 5 times less effective of a protein than canine IL-4.  In the first round of experiments, I had been putting an amount of IL-4 into the wells equivalent to that of the dog experiment (2ml IL4/ml of media). However, if the equine protein is less effective, a much higher concentration of IL-4 must be added to the cells in order to induce a similar response.  We also decided on a new formula for growth media, consisting of horse serum as opposed to the original fetal calf serum, and pen strep as an antibiotic as opposed to what we used the first time (gentamycin), for there are published papers about people culturing equine B Cells using pen strep.

We are also adding a component to this experiment.  Some of the B Cells are going to be CFSE labeled.  This will tell us if cells are not proliferating at all, or if they divide and then die off immediately.  CFSE dies the cells a dark green and with each division of the cells the intensity of the green is cut in half.  Therefore we can flow the cells and the machine will tell us how intense the green is in relation to that very dark shade.  This will tell us if we are not getting any proliferation at all, or conversely, is the cells proliferate but their living conditions are such that the cells cannot survive.

New blood arrived on the Monday of my last week and again the PBMCs were again isolated and plated with and without feeder cells expressing CD40L.  They have been cultured with the new growth media and received a more concentrated dose of IL-4 so we will see how this one goes!

The Mason lab is embarking on a new project. Led by the PhD student Kazim, the first infusion of CAR-T Cells will be infused into a German S

heppard with lymphoma named Lady.  The CAR T Cell project is really remarkable as it uses the HIV Virus to infect healthy T-Cells that become “serial killer cells” that seek out and kill tumor cells.  Carl June, a revolutionary researcher at Penn, has done some incredible work with CAR T cells and you should really watch this video if you have 5 minutes. Its so cool.

https://www.youtube.com/watch?v=h6SzI2ZfPd4

Ms. Cozine came to visit me last week which I thought was pretty funny because the last time she saw me in a science setting was when I was in her freshmen chem honors class! I taught her how to count cells via hemacytometry, we made LB Agar plates, and even made some more 70% EtOH (ethanol) solution before meeting everyone for lunch.

 

Hemacytometry -- a 10ul sample of the cells in their growth
media is put in this little glass frame containing a tiny tiny grid that looks
like this under the microscope--you count all the cells within the boxes and
plug that (when multiplied by the volume of the media in the vial)
number into a big equation that gives you the total cell count in
the media.

A picture of my building I snagged on my way home on my second to last day.

Wednesday, my last day, was incredibly sad.  We all went out to dinner at Mad Mex and my friends from the lab gave me presents, including a necklace made from the exact vial from the liquid nitrogen freezer that once contained a type of cell that I mistakenly thawed and cultured.  Pretty funny.  Everyone has tried to convince me to go to Veterinary School while I have been here and they even gave me a Penn Vet T-shirt.  EXP has easily been the coolest experience I have had in any summer and I could not have asked for better people to work with, a cooler topic to study, or cooler city to do EXP in! From Phillies games to Cheesesteaks to the Palestra, and meeting my fellow EXPers in the city, it was all pretty cool.See you all in September!

Met Bridgid for a Phillies game.