Week Eight . . . The End

After eight weeks of living and working in Cambridge, MA, my time here has come to an end. My last few days at Harvard and MIT were quite eventful, so much in fact, that I slept a total of less than 14 hours during my last three nights! But I’ll start at the beginning . . .

On Monday, I spent nearly five hours extruding in the lab. Extrusion, if you remember from my last blog entry, is one method of forming liposomes of a desired size. The method is relatively simple; the solution containing the liposomes is filtered through screens of decreasing size using high pressure. The solution needs to be filtered through the larger filters three times, but through the smaller filters 12 times. So the process takes a long time when there are six filters involved. Also, in between each filtering cycle, the extruder needs to be disassembled and cleaned for any excess solution. I was standing during the whole process, so I was pretty tired by the end of the day.

On Tuesday, there were more presentations from college students in journal club. Topics included the creation of gels held in dialysis tubing for controlled release drug delivery and the development and evaluation of 3D gel constructs for vocal fold tissue engineering. Another student presented a paper on the dispersion of biofilms with engineered enzymatic bacteriophage. I enjoyed the two student research presentations, but had some difficulty with the content of the paper. Back in the lab, I weighed out synthetic lipids and cholesterol for Al and Hila’s future work with the liposomes.

Wednesday was my last day in the lab, and I spent it working alongside Al and Hila, freeze-thawing four different liposome compositions. Three of the samples had the toxic substance saxitoxin (STX) inside of them. STX is a naturally-occurring toxin that is a potent and extremely selective sodium-channel blocker. It is responsible for red tide and the subsequent harsh effects on humans. In fact, ingestion of 0.2 mg of STX is fatal to an average size human, so I was a little wary in handling the substance, even if it was safely enclosed in a plastic test tube. Other than the freeze-thawing, there was little for me to do around the lab, so I spent some time cleaning and saying good-byes before I left.

Meanwhile on campus, it was finals week for everyone. Monday night, I was up until 3:00 studying for my Tuesday exam. After only a few hours of sleep that night, Hila and Al gave me strict instructions to take a half day at the lab and go take a nap before my exam. The exam itself was okay. I was more relieved than anything after it was over, which my whole floor heard about while I danced in the hallway.

Wednesday was the last day on campus for most of my floormates, including myself. To celebrate, we had a gigantic sleepover (involving very little sleep), complete with 15 cartons of strawberries and more food than we could have ever possibly eaten. It was a fun way to spend my last night, even though I was scrambling to finish my packing on Thursday morning before my parents picked me up. The morning concluded with my last breakfast in Annenberg and even more goodbyes in my dorm before the ride home.

All in all, these last eight weeks have been pretty amazing. What I’ve learned most about research is that you can never predict how busy a lab will be and you should always prepare for the worst-case scenario. As for myself, I learned how to survive away from home and challenge myself in a class where it was impossible to know all the answers. Thank you to Mr. Palm, the Science Department at Brooks School, Harvard Summer School, and MIT’s Langer Lab for giving me the opportunity to have such a once-in-a-lifetime experience. I encourage other Brooks students to apply for future summer opportunities.

Have a great school year!

Week Seven . . . A Little Bit of This, A Little Bit of That

Seven weeks down, one to go. It’s scary to think that I have less than a week left in Cambridge. As far as busy weeks go, this was one of the busiest. I spent my week shadowing Al and Hila, mostly observing and learning about their work with liposomes.

Liposomes are another type of particles used in experimenting with drug delivery. The basic process of making liposomes, as Al explained to me, is as follows:

First, all the hydrophobic components (lipids, cholesterol) are dissolved in a chloroform: methanol solution. Then, the chloroform and methanol are evaporated, using a rotovap at a temperature above the melting point of the lipids. A rotovap is a machine that uses temperature and pressure as a means to evaporate and then remove liquids from a solution. When the chloroform and methanol are evaporated, all that is left is a film. An inorganic substance called t-butanol is used to dissolve the film, then the solution is frozen in liquid nitrogen. After 2-3 hours, the t-butanol is evaporated using the rotovap again and the remaining film is put on the lyopholizer overnight. Then, you add water and ammonium sulfide to return the frozen sample to an aqueous phase. At this point, the process for making liposomes diverges, depending on the size you desire. Extrusion, or filtering, is one method. The other method is to homogenize the sample for 10 mins, adjusting the speed based on the desired size for the liposomes. Then, the sample is freeze-thawed (transferred back and forth 12 times from liquid nitrogen to hot water). No matter which method is chosen, the final steps include dialysis to get rid of the ammonium sulfide, loading of the drug, and then dialysis to get rid of any outside remnant drug.

As you can see, making liposomes is a long process, but I was able to see the entire process at different points throughout the past week.

In addition to the liposome work, I worked with Al to take pictures of embryonic stem cells in some of the HA hydrogels that Sarah and I had made. I also watched Hila perform a Live/Dead assay on the same cells. I finished the MTT assay for Sarah’s SBA-15 experiment, and my results showed that the supernatant alone had little to no effect on cell viability.

On Tuesday, and extra-early journal club consisted mostly of presentations from summer interns preparing to head back to their respective universities. Topics of discussion included targeted gene delivery of nanoparticles to MDA breast cancer cells and the creation of ‘patterned monolith structures for passive mixing in microfluidic channels.’ Later in the week I attended a poster session for other summer students, where they were able to present their research for the MIT community. I learned a lot, and I was surprised at how much I understood. Also at journal club, a graduate student presented a paper on continuous sensing with magnetic nanoswitches.

Wednesday, I met my aunt (who works only a street away from Langer Lab!) for lunch and we did some shopping in between my liposome work at the lab. Thursday was the Langer Lab beach party. However, because I had my last lecture of the summer session that night, I didn’t attend the party, but instead went on a college visit with my parents before coming back just in time for class. Our last lecture topics included the sexual transmission of HIV, development of a vaccine, and HIV in India.

This weekend I “visited” Harvard with my parents, attending an information session, but opting out of a tour, since I’ve seen nearly all of campus while living there. My final exam is Tuesday night at 6:15 pm, so I spent the remainder of my weekend studying anywhere and everywhere: my room, the library, the science center, the lawn in front of my dorm, the tutoring office, and the hallway. I did, however, make some time to support Jee Su at an orchestra performance and stop by the last dance of the summer.

With only three more days for me at the lab, a final exam, and all my packing to do before I leave on Thursday morning, these last few days in Cambridge will be stressful and busy as everything comes to a close. Wish me luck!

Week Six . . . Polymers, Buses, and YouTube—Oh My!

With my sixth week in Cambridge now behind me, I’m my time here is quickly ending. This was my last week with Sarah as my post-doctorate, and we spent most of the week finishing up things so that she could get ready for her trip to Canada. We did have a decent amount of lab work, but compared to some past weeks, it was much quieter and more relaxed around the lab.

This week we focused on polymers, a stark contrast to MTT assays. We made a PXC-ADH polymer again, following the same process of maintaining the pH while we added the materials, putting it on dialysis and then freeze-drying it. Later in the week, we dissolved the freeze-dried PXC-ADH along with several other polymers. We tried mixing different combinations of the polymers at different concentrations with a double-barreled syringe to see if any hydrogels formed. This process took most of the week, so I spent time cleaning up the lab bench and reading scientific journals as well.

For this week’s journal club, there were talks on the ‘disassembly of nanodiscs with cholate’ and the production of neural-controlled prosthetic devices for humans with tetraplegia. The first talk was way over my head, but I found the second topic quite interesting, especially the videos we were able to see of the working devices.

On Thursday, I plated mesothelial cells on my own for the first time. Dan Anderson, the head of our group at MIT, asked Sarah to perform a rough experiment on the effects of SBA-15 nanoparticles versus SBA-15 supernatant on mesothelial cells. Since Sarah won’t be around to finish the experiment next week, I was responsible for adding the substances to the cells on Friday and I will perform an MTT assay on them to determine what happened to the cells in the different environments.

In class this week, we discussed the human immune system: how it reacts to HIV infection and how HIV turns the immune system against itself during progression to AIDS. It was some of the hardest biology we’ve discussed so far; I even had to pull out my AP Biology book to review basic immune responses so that I could understand some of the more advanced concepts.

Friday night, Jon and I had our first adventure on the bus system in Cambridge, venturing to Mr. and Mrs. McCahill’s new home for dinner. We spent a few hours with them before heading back to campus. I was also home for part of the weekend, spending some time with friends and working on one of my community service projects for this school year.

Sunday night I received an urgent text message from a fellow Brooksian instructing me to return to campus in time for the Secondary School Talent Show at Harvard. So after gathering all my stuff in less than ten minutes and suddenly informing my parents that I had to go back to campus two hours earlier than anticipated, I was there to see five old and new friends perform—to a Backstreet Boys song. It was hilarious; they’ve now become campus celebrities and videos of their performance can already be found on YouTube. My weekend ended with an 11:30 pm ice cream run with a few friends who we convinced to stop by Harvard Square on their way home from Boston.

Next week I’m going to be working with Hila, a post-doc, and Al, a student at MIT. I’ll be shadowing the two of them and their work with liposomes in addition to completing Sarah’s experiment. I’ll be seeing a multitude of different things in the lab, so it will be a change from my current routine.

Enjoy your week!

Week Five . . . A Focus on MTT Assays

My fifth week in Cambridge has just finished and I’m finally getting the hang of things. This week I worked mostly on my own in the lab; Sarah gave me a list of responsibilities for the week, broken down by day. I had to be efficient so that I could get everything done each day, and I also had to pay close attention to the timing for the different steps of experiment protocol.

This was MTT week for me, meaning that I performed a lot of MTT assays. As explained in a previous entry, an MTT assay is a colorimetric test that observes cell viability in a variety of different environments. Sarah and I tested the viability of cells exposed to different nanoparticles. Monday morning I added nanoparticles to a few well plates of macrophages and mesothelial cells. Then I performed an entire assay on plates of C2C12 cells that Sarah had added particles to over the weekend. Each plate had a different concentration of nanoparticles in a buffer solution, so we were able to observe how that affect cell viability as well.

Tuesday morning started with Journal Club, where we heard presentations on detecting proteins and engineering vascularized muscle tissue. Then I was back in the lab performing MTT assays on another batch of mesothelial cells and macrophages. Tuesday afternoon I created Excel spreadsheets with our growing data so that we could look for trends over time. Tuesday night was my midterm exam, a two hour test that I spent Sunday night, all afternoon and night Monday, and most of Tuesday afternoon studying for. It was hard, but I think I did okay.

Wednesday I did more MTT assays and watched Sarah do a Live/Dead assay on the same nanoparticles I was testing. I also caught up on some reading for class. Wednesday night Jon, Jee-Su, and I met Mr. and Mrs. Palm for dinner in Boston. After going the long way around Boston Common (but never actually getting lost), we made it in one piece and had a great meal, catching up and sharing stories from around our labs. That night, we also had a study break on my floor of Hurlbut, where we ate even more and met the second session students that moved into our dorm for the last four weeks of the program.

On Thursday the MTT assays continued, and our data really started to take shape. I was able to create graphs for the C2C12 cells. By this point we had finished assays for all of the C2C12 cells so we had a complete set of data. This weekend, Sarah is going to write up a summary of what we’ve done and the results of our assays so that she can use it in a draft of a paper she’s starting. Thursday evening, I had my discussion section and then stopped by Annenberg Dining Hall to eat a Harry Potter-themed meal, complete with a high table, teachers in costume, and British cuisine. Then it was time for lecture, where we began learning the biology of how HIV is transmitted and how it damages the immune system so effectively.

I was on my own in the lab on Friday, where I did the last MTT assay of this set and cleaned up the 20+ well plates that we used this week. I also changed the water for the polymer we had dialyzing. Next week will be a “polymer week,” according to Sarah, with focus on creating better hydrogels that we did last week.

My sister came down Friday to spend a night with me at Harvard; we went out for dinner and a movie and explored Harvard Square. Saturday we went out for breakfast and walked along the Charles before going home to the air conditioning and escaping the heat of Cambridge.
This coming week is Sarah’s last week with me before she leaves on a two-week vacation. I’m going to be working under Hila, another post-doc, for most of my last two weeks. This week I’m going to be introduced to some of her projects so we can determine what I will be working on during that time.

As usual, have a great week!

Week Four . . . Halfway There!

It’s hard to believe that my time at Harvard and MIT is already halfway over. With four weeks down and four weeks to go, this week marks the midpoint of an amazing summer.

Contrary to my last entry’s tone, this week was anything but slow at the lab right from the start. On Monday, with two different polymers dialyzing, I was constantly checking my watch to make sure I didn’t miss changing the water at the correct time. We also created another polymer, which we left stirring on a hot plate overnight after adding various substances that needed to dissolve.

Journal club met as usual on Tuesday morning. This week’s presentations addressed how to use peptides to cross the bloodbrain barrier in order to distribute medicines (a topic also discussed in my HIV/AIDS class this week!) and how Markov chains can be used to predict the outcome of baseball games. After journal club, I changed the medium for all 10 plates of C2C12 cells and checked them under the microscope. After changing the water a few more times, I took the polymers off dialysis, freeze-dried them, and put them on the lyopholizer so that the water could be vacuumed out. Tuesday I was invited to attend a lunch with Dr. Langer and the other undergraduates and high-schoolers interning at the labs. We were able to ask him a variety of questions and congratulate him on his most recent accomplishment: being awarded the National Gold Medal of Science.

On Wednesday morning, Sarah and I defrosted macrophages and mesothelial cells and began growing them in flasks. Next week, we’ll transfer them to well plates so that I can perform MTT assays on all of our cells: macrophages, mesothelials, and C2C12s. We made yet another PXC polymer, which involved constant monitoring of the pH, before we put it on dialysis. For lunch on Wednesday, I took the train to Charles/MGH and ate with my uncle. I haven’t gotten to explore too much of Boston, so this was a new experience.

Thursday I changed the water for dialysis before our weekly Kohane group meeting. It went pretty long, so we didn’t spend any time in the lab in the morning and I caught up on some reading for class instead. In the afternoon, I saw the cold room, where we store a majority of our liquid supplies, for the first time. Sarah and I made mesothelial cell medium, a long process with many additives, including insulin, which I had to put into tiny 275 containers to store for another time. We also started making the nanoparticles to add to our cells for our experiments next week.

On Friday I changed the medium for the C2C12 cells again; Sarah will add the nanoparticles to them this weekend, and I’ll do the MTT assays on my own next week. We took two of our polymers off dialysis and experimented with different reactions, using a double-barreled syringe to see if we could make hydrogels that can be injected into the body. I also flew solo with the infamous lyopholizer (see photo below) and put some of our samples on there to dry.

It was Harry Potter weekend at Harvard. And of course, being an avid fan, I was in line to get my copy of the book at 12:01 am on Saturday at Queen’s Head Pub, a student hangout below Annenberg Dining Hall. It was a weekend of reading on campus, both Harry Potter and my AIDS textbook, as I prepared for my midterm exam, which is on Tuesday the 24th. However, I still found time to socialize; a few friends from Brooks came down to visit on Sunday for the day. I also had dinner in Cambridge with my family on Sunday night. This coming week will be busy, with my midterm studying, a full schedule at the lab, and a dinner planned for all the Cambridge interns and Mr. Palm.

Have a great week!

Week Three . . . Never a (Really) Dull Moment

Lab work is never steady. On the one hand, there are bursts of time when I am running around trying to plate cells and my thumb feels like it will fall off if I even see another pipette. On the other hand, there are days when I sit around reading scientific journals, waiting for something to do. This was the lesson I learned during my third week at Langer Lab.

Monday morning, I changed the water for dialysis once more. Then Sarah and I poured the polymer solution into tubes that we centrifuged and then freeze-dried, so that the water could be vacuumed out. To freeze-dry them, we placed our samples in liquid nitrogen. Then the samples were put in a glass flask and into a lyopholizer, where the solvent is removed by vacuum over the course of a few days. That afternoon, I performed an MTT assay on my own, adding dye and stopping solution to macrophages and mesothelial cells, then incubating them for the night.

Tuesday morning I attended Langer Lab’s weekly summer journal club meeting, where different researchers (post-doctorates, graduate students, and undergraduates alike) present scientific journal articles that they’ve recently read while we have a group breakfast. This week’s articles were about the new HPV vaccine and DNA folding. Later that day, I changed the medium for our C2C12 muscle cells. Sarah also taught me how to store our absorbance data from the MTT assays in Excel spreadsheets and graph the data so we can track cell viability over time.

On Wednesday, things slowed down around the lab. In the morning, we had a Kohane group meeting, where we heard about current and future experiments from each researcher. Wednesday afternoon, Sarah and I took a walk to the Histology Department to drop off some dissection samples to be analyzed.

Thursday was proof that no matter how much you plan, things will still go wrong. After nine long days of growth and incubation, we discovered that our C2C12 muscle cells had all died, suddenly clearing our schedule for the day. Then we discovered that the lyopholizer had been used improperly and all the samples on it had melted, including the polymer that we had started making a week and a half ago. We were able to save some samples, but we still lost a lot. After a few hours of reading journals and studying for class, we went with another researcher to an NMR (nuclear magnetic resonance) machine and analyzed some sugars.

Friday the 13th was actually a much luckier day than Thursday. We were able to plate 10 sets of C2C12 cells that will grow for the next nine days. We also started making a new PXC-M polymer; to finish it will take all of next week and perhaps even some time in the following week.

There were good days, bad days, busy days, and even a few quiet days in the lab this week. Back at Harvard though, things were pretty busy. I spent most of my evenings reviewing for class and catching up on my summer reading. Class is going well; I spent some time at Brain Break on Tuesday night with the tutor for my section, reviewing while we drank hot chocolate and ate cookies. After 3 ½ hours of class on Thursday evening I was pretty tired, but I still found myself eating ice cream in the dorm at 12:30 am with some friends.

This weekend I’m bringing a few Brooks friends back to my house for some well-deserved homecooked meals and relaxation. Sunday we’ll be back, ready for whatever week four will bring.

Have a great week!

Week Two . . . Cells, Cells, Cells

My second week in Cambridge flew by. Finally, I was able to get more involved and even work on a few different experiments. On Monday, Sarah and I created a polymer that will be used to make a hydrogel next week. My role was to constantly monitor the pH level of the mixture, maintaining it as close to 6.8 as possible. I measured the pH after we added in each substance and used NaOH to raise it back to 6.8 if it dropped. After it was stable, we left the mixture stirring for a few days.

Monday afternoon, I went with Sarah across MIT’s campus to work with TEM (Transmission Electron Microscopy). MIT has four of these huge microscopes, which Sarah used to take pictures of the microscopic channels in three different particles that we’re going to work with early next week.

Tuesday was a busy day for me in the lab. In the morning, I learned how to do an MTT assay, which is a laboratory test that uses color to measure cell growth. Basically, we culture cells, transfer them to well plates, add medium, change the medium every three days, add the materials we’re testing and then stain them with a dye to see if the cells are still alive. It’s a simple process, but time consuming. After observing, I was able to change the medium for a few samples and even fill the well plates. By the end of next week, I should be able to perform nearly the entire assay on my own.

In the afternoon, I went with Iris, another researcher, to observe her work with animals. She practiced making nerve blocks on the cystic nerves of rats. The process involved injecting lab rats with a mixture that would “block” the nerve, or make it temporarily non-functioning. Then she went through a series of tests to determine if the nerve was actually blocked. After being layered in a coat, hair cover, mask, gloves, and shoe covers, I wrote down the results of the tests for her. It was interesting to see her work with the rats because I’m learning that animal work is an inevitable part of biological research.

I had the day off on Wednesday for the holiday, so I spent the day at home with my family.
Thursday I spent most of the morning reading about different assays that we will do during the next six weeks. In the afternoon, I learned how to count cells, a somewhat tedious process, and how to determine if there were enough cells in a culture to perform an experiment.

On Friday we put our polymer into dialysis bags and suspended the bags in water. It was my responsibility to change the water every four hours. Next week, we should be able to freeze the mixture in liquid nitrogen and vacuum the water out, so we can start making the gel. I also changed the medium for C2C12 (muscle) cells, which involved vacuuming up the medium and pipetting fresh medium into 96 well plates.

So after a busy week, I’m heading home for the weekend, to spend some time with my family. On Sunday evening, I’ll be back for a dormwide cookout and a meeting with the tutor for my HIV/AIDS class. Have a great week!