WISE journal entry # 17

Name: Leah Gerald

Entry: #17 (4/18/2012)

WISE Week: 8

Written on Date:4/23/2012

Mentor: Sagar


today my mentor planned to stain the mice brain cells we worked with yesterday using immunoflourescence. this type of staining helps show human cells, and since the cancer they inserted in the mice was human based cancer, it would make the cancer cells stand out. unfortunately, I wasn't around for this part, but i was around when they performed the transfection for another experiment. they added some plasmids to some gbm (glialblastoma) cells they had been working on for a while. next, they needed some cells to be re-seeded into a smaller plate for them to photograph their movements. these containers they put them in contained very small, controlled areas that they would put under the microscope with a camera and photograph its movements over time to see how it migrates.

WISE journal entry # 16

Name: Leah Gerald

Entry: #16 (4/17/2012)

WISE Week: 8

Written on Date:4/23/2012

Mentor: Sagar


today was the day that a large accumulation of my mentors works came together. we looked at the brain slices they had taken from the mice they injected tumors in after months of preparation. first they had to accumulate and culture the cells, then once they had enough, they had to inject them into the mice, then, BEFORE the mice died, they had to extract the brains and freeze them. then another member of the lab had to take those mice brains and meticulously slice them into tiny slices to put under the microscope. then they had to add different types of staining, which they had to deduce which type of staining they used (different for different cell types) after all of this, they had to then sort the sections of the brain they took into slides with cancerous cells on them, slides without, and find the slides where the point of injection is obvious, because that area should have the largest concentration of cancer cells, and also, it helps as a reference point. ONLY after all this can they make deductions about the cells they are working with, and even then, they weren't fully sure of everything. there were sections of the brain that looked very strange to me, so I asked my mentor about them, but they didn't know what they were. so there are still things they do not know.

WISE journal entry #15

Name: Leah Gerald

Entry: #15 (4/11/2012)

WISE Week: 7

Written on Date:4/11/2012

Mentor: Sagar


today went by very fast because I was able to do a lot in the hood. we started out by checking on the pancreatic cells I set yesterday. Sagar said they looked very good, and then I got a chance to look at them

they looked very healthy, and somewhat more different from all the other cells they culture, but because they express that common gene (brachiary) they share at least one similarity. after checking on this, they needed to put some new media in a large amount of the Cordoma cells, and they needed to culture some other cells because they had become too confluent. first, I aspirated all the containers and then pip-petted the new media into each container. it was a long process because there were a lot that needed new media, and as I did this, I didn't realize how much time had passed. the process was very menial and boring, but time just flew by. next, there were some cells that were getting cramped in their container, so we aspirated the media, added trypsan so the cells would un-stick from the bottom of the plate, spun down the cells, mixed them with new media, and re-plated them. the process is one I have watched done several times, except this time, I was able to take part in it as well. at this point, it was time to go, so my day was done.

WISE journal entry #14

Name: Leah Gerald

Entry: #14 (4/10/2012)

WISE Week: 7

Written on Date:4/10/2012

Mentor: Sagar


today, we were using the western blots previously formed to make one of these:

in order to produce these images, we have to go through a photo-like procedure. first, I washed the membranes in a buffer three times to make sure they were clean. then, we mixed in rabbit antibodies and mouse antibodies, one in each of the trays that corresponded with whichever type of cells. we let those sit for a little while, and while we did that, we went upstairs to get some pancreatic cancer cells. my mentors had found out that there is a gene, Brachiary, seperate from the hippo gland, that can also have an affect on YAP proteins. they found this gene was present in Cordoba cells, which they have in the lab, but recently, they found out that it is also present in pancreatic, colon, and some forms of lung cancer. because of this they were hoping to begin to observe this gene in action in the other types of cells, so I cultured the pancreatic cells into its proper media, and stored it with the others. after this, the western blots were ready to be developed. we poured out the antibodies, and washed them in a sort of photo-developing chemical that brings out light where proteins are present when developed. we then went into another lab, where there is a room to develop westerns, placed them in a machine that does all the developing work for you, and make a couple of images to compare the proteins. at this point in time, I was late for pickup, and I had to go.

WISE journal entry #13

Name: Leah Gerald

Entry: #13 (4/4/2012)

WISE Week: 6

Written on Date:4/10/2012

Mentor: Sagar


today I was able to fully witness, until the last step of course, the process of making a western block. a western block is something my mentors use to compare the amount of proteins given off by certain cells in certain conditions.

the different rows (horizontal) show how much protein appears in different conditions. the lines (vertical) show the different sizes of proteins and where in the spectrum your protein of interest falls.what we had to do first, was mix the protein with a dye so it would be easy to see. then we had to put the protein in a hot bath, because proteins take a string-like shape, and when they are in the mixture they are all jumbled up. heating them up makes them spread out somewhat and become un-jumbled. after that, they are poured into a gel, which is in a container that can send an electrical current through the gel.
once all the proteins are sorted into their specific compartments in the gel, we then turn on the electricity, and the lighter proteins travel down the gel through the current. after about an hour, the protein has shifted all the way to the bottom, and at this point, my time is up, and I must go once again.

WISE journal entry #12

Name: Leah Gerald

Entry: #12 (4/3/2012)

WISE Week: 6

Written on Date:4/3/2012

Mentor: Sagar


today was an exciting day because I finally got to work under the hood! the main things that they had to get done today was, Sagar needed to do an RNA synthesis, but first, we needed to culture some cells. the breast cancer and breast cells from a while back were becoming confluent (basically there were too many in one container) and they needed to move some from one container to another so the original ones would have room to grow and expand. because it was a task that didn't require too much precision, Sagar let me work with him under the hood. to start with this process, we sucked all the existing media out of the containers. then we added an enzyme called trypsin, which makes the cells disconnect from the base to the flasks. with that done, we shook the containers and made sure all the cells disconnected before we added calcium, which acts as an inhibitor, and stops the trypsin from acting on the cells too much. next, we took the cells in the flasks, and put them in tubes. we then put the tubes in the centrifuge and spun them until all the cells were in a pellet at the bottom. we took the tubes back to the hood and sucked all the trypsin and calcium out of the tubes. once they were all without trypsin, we filled the tubes with media and mixed the cells with the media.we then got new flasks, and put part of the cells in the tubes in the flask, along with some new media and put them back in the incubator.

WISE Journal entry #11

Name: Leah Gerald

Entry: #11 (3/28/2012)

WISE Week: 6

Written on Date:4/3/2012

Mentor: Nathaniel


today was different because Sagar had to take a test, so he was not at the lab today. Nate acted as my mentor in his place for a day. there wasn't much to be done, because they hadn't been able to do much recently, so Nate took me to the hood to feed the cells new media. basically, all of the cells they culture live in little bottles filled with a substance that contains the nutrients they need to survive. as the cells eat up the nutrients, the pH levels in their food change, and they change from red to yellow. Nate took out all the cell cultures that were yellowish, aspirated out the food, and refilled it with more media. Its OK for them to suck out the substance like so because the cells are attached to the bottom of the plate, and when the new media is added, they were hungry for only a short time so they survive. on another note, I finally met Dr. Q today. he didn't have much time, and it was truly a coincidence he was here, but I saw him, shook his hand and said a few words, and then he was off again, taking care of some crises in the research lab. that was the highlight of my day.

WISE Journal entry #10

Name: Leah Gerald

Entry: #10 (3/14/2012)

WISE Week: 5

Written on Date:4/3/2012

Mentor: Sagar


Today was a roughly slow day, because once again , there was nothing yet for me to get my hands on and "do" but rather a lot of watching to be done. Nate was working on making a control comparison of the DNA/RNA levels in a sample of some proteins they had been working with. To make this control, he first wanted to find out the concentration of plasmids (the shape DNA takes for bacteria. like a circle instead of the double helix) that is he needs to get an average concentration, which is 230/260. He tested them in the protein assay machine, and found out the amounts he needed of 3 different strands: one was the first part of the plasmid, then the second part, and then the full strand of the two together. This way he could see different areas of expression in the genes. after this, I watched as Sagar transfected the cells in the hood. He used the plasmids Nate had set up earlier to transfect the cells. Its hard for the plasmids to bind with the cancer cells, because plasmids take on a circular shape, and the cancers DNA is in the double-helix shape, but overtime it does bond and starts to express whatever genes were in the plasmids combined with its own. and with this, I had to leave, and my day was over