January 7th - January 31st
Unfortunately for me, I was unable to do much of anything in terms of hands-on research during the month of January. My on-boarding process was backed up and my criminal background check was lost in the MGH system. That being said, I was still able to meet on a regular basis with Dr. Castro and the post-doc I am working directly with, Hyungsoon Im. During these meetings, we discussed where my focus should be during my time with CSB. Essentially, it was boiled down to two main projects. First, I would help Hyungsoon process patient, cell line, and mouse exosome samples from various cancer types so as to validate a new technique developed by Hyungsoon and others at CSB. We would utilize a recently developed nanosensing device called the nanoplasmonic exosome assay (nPLEX). I spent about a week of this time on my own delving into the literature on this novel platform so that when I began my research I could hit the ground running.
The second project would involve the nPLEX again, but this time it would be used as a validation tool for the antibodies used in each experiment. A big problem in our lab is that the same antibodies received from the same vendor often yield inconsistent results in lab testing. That being said, my second goal for this co-op period is to develop a protocol in which the antibodies can be properly validated so that our lab can utilize the most effective ones for our experiments in the future.
In addition to project planning, I was able to begin sitting in on weekly journal club, BME, and large group meetings with our head PI Dr. Ralph Weissleder. These are excellent learning tools, not only for content but also for how to best present my work to others in the scientific community. I learned so much about the research being done within CSB and elsewhere in the world, and it is truly remarkable and eye-opening.
February 1st - February 14th
The first two weeks of February were more of the same in terms of being cleared by MGH. I still did not have a badge, but I began to visit the Charlestown Navy Yard Building where I would be doing most of my work. I was able to shadow Hyungsoon through a full nPLEX protocol and he was very helpful in answering all of my questions. In addition, I learned how to utilize the data collection program for the nPLEX and the microarray spotting system which will prove to be a huge part of my co-op experience. That being said, nothing very new and exciting happened during these two weeks.
February 16th - February 28th
Good news! I was finally cleared and received my badge from MGH! So it was only a month and a half late. These two weeks were spent with the microarray spotting system learning how to create programs for nPLEX alignment validation. The nPLEX chip is mounted on a glass slide, so we wanted to develop a program so that each nPLEX could be placed on the same location of each glass slide. Having the nPLEX in the same location will prove to be very important for experiments down the road. After several days of wrapping my head around the microarray spotting technology, I developed a pretty simple program to spot four drops of DI water, one on each of the four corners of the chip right next to a 1mm x 1mm hole marker. If all of the spots were visible, then the chip was well aligned and could be used in another experiment. Essentially, this period of time was useful as a warm-up for the work to come in my project.
March 1st - March 27th
Building upon some of my experience with the microarray spotter, I was tasked with creating a program that could spot 25 different solutions onto 100 arrays in a 2x2 array pattern. We wanted to establish this program so that we could conjugate up to 25 different antibodies to be run against exosome samples. In addition, instead of using all 25 solutions, we could use 12 antibody solutions and run two exosome samples on the same nPLEX chip. This would increase our throughput for the coming experiments, and greatly reduce processing time for an equivalent amount of samples. With the help of Hyungsoon and the occasional troubleshooting with a specialist from the microarray spotting company, we were able to successfully create a program that provided our desired results.
That being said, it took quite some time to optimize the conditions of this program. At first, we ran into quite a few issues with the microarray spotter dispensing tips. These had not been used for several months, and two of them had been partially damaged in previous program test runs. We attempted to use the tips in our new program, but unfortunately they were not capable of dispensing the proper or constant amounts of solution. Hyungsoon and I ordered two new tips from the company and after a week they arrived. Once we had removed the remnants of the old tips and inserted the new ones, we ran the program through several times. Though the drops were now uniform in size, each tip was dispensing solution at slightly different coordinates even though the program coordinates were left unchanged. We tried a lot of different possible solutions to this problem and eventually we were able to correct the issue. Now we were ready to try our nPLEX improvements on real experimental data!
March 30th - May 1st
I spent this month honing my skills on the nPLEX protocol. I started by running a few glioblastoma mouse samples against a set of 12 different antibodies. Instead of using 25 different markers, we decided to use 12 in duplicate across the chip so that we could run 2 exosome samples at one time. My results steadily improved as I learned that I needed to be on my game through the entire protocol, paying attention to every minor detail so that I wouldn't skew my data. As I became more accustomed to the nPLEX protocol, we moved into preliminary data collection on our proof-of-concept study using pancreatic cancer patient samples. I began running some healthy controls, and once the data correlated well with expected results, we moved on to the pancreatic cancer samples.
May 4th - June 1st
The proof of concept study began to take up most of my time at work during this month. There was a grant deadline that on June 1st that we needed to run 30 patient samples for in order to secure further funding. About two weeks into May, I began to run 2 chips a day, meaning 4 patient samples per day to ensure the data would be collected by the first of June. It was certainly a very stressful time, but I organized my time well enough to complete all the necessary samples: 25 pancreatic cancer samples and 5 healthy controls. It was a rewarding experience to complete this task as part of a real research team; it really put into perspective how lucky I am to have secured a CaNCURE position. In addition to the stress of running samples night and day in the month of May, I was assigned a journal club presentation for the end of the month. I presented a paper from Nature Medicine titled "Studying clonal dynamics in response to cancer therapy using high-complexity barcoding." The presentation was in front of approximately 15 post-docs from the Center for Systems Biology. I will be honest, it was a very nerve-wracking experience. I am happy I accepted the challenge though because it made me so much more comfortable presenting in front of an audience. It was interesting having to defend someone else's research methods, findings, etc. as opposed to mine for a change. Overall, I'd say this month was the busiest of my co-op experience.
June 1st - June 26th
After having collected the data from 30 samples, we paused for a bit on the proof of concept study to accumulate more patient samples and direct our research in a more pointed way. That being said, the month of June slowed down a bit in terms of research production. I began work on an antibody validation study to determine how quickly antibodies degrade when stored at varying conditions. This data is still in process as we have had a few issues lately with the analysis portion of the protocol. I worked on my CaNCURE Day poster quite a bit, and likewise prepared a final co-op presentation summarizing all of my work that I have completed this semester. Putting all those things down into poster/presentation form really opened my eyes to how much I contributed to my research team. As my final task before leaving my full time position at the lab, I was instructed to train a new summer student on the nPLEX protocol so that he could continue work similar to what I did in the future.