Week 7: Confirmatory Western, ELISA - Alexandra Kot

Alexandra Kot | Dr. Sandra Demaria 


2026.07.13:

After growing my suspected MAVS KO TS/A cells over the weekend with a control TS/A, they needed to be irradiated to activate MAVS production. The cells underwent 3 minutes of irradiation, for a total delivery of 8 Gy. In two days, the cells will be processed again for a Western blot. 

2026.07.14:

Today was the first day of clinical immersion presentations, with most presentations coming from the bone-centric and computational students. It was really interesting to hear about Manav's presentation, focusing on graft deterioration over the extensive testing period required before implantation. Manav looked at both life-dead cell staining and mitochondria depolarization. Pooja's alloplastic ear reconstruction project was also very interesting. I hope the Spector lab can use her work to make an ear replacement. 

I was able to catch the second half of the Demaria lab meeting and listened to Dr. Elisa's presentation. She is a visiting postdoc who worked on cell-inspired microrobots, which are pH-responsive and ultradeformable. 

With my successful MLKL knockouts, the next step is to perform an ELISA to quantify how the gene knockouts could affect interferon production. Yesterday I seeded the knockout MLKL cells from #13 and #14, so today I collected the media for the proliferative cell samples. The interferon produced by the cells is excreted into the media. The resulting cells underwent 8 Gy of radiation. Every day, until Saturday, I will sample the media from the cells to capture interferon production in the days after the cells are irradiated. The concentration of interferon in the media will be normalized to the cell population. 

2026.07.15:

Today was my first operating room day. I shadowed Dr. Spector with the help of Justin and Pooja. There were three surgeries today. The first, which freaked me out the most just based on novelty and the amount that the cauterizing wand was used, was a breast reduction. This patient had four doctors working on her simultaneously, and they worked with great precision and grace. It was really nice to see. I always had the image of the OR being a very hectic and dramatic place (probably not helped by medical dramas). The second surgery was a lip graft for a burn patient, where Dr. Spector cut a portion of the lip that had fused to the patient's skin below her lip to release tension. After which, the expanded wound was replaced with a skin graft to prevent further tension in the healing process. This was the quickest surgery. The third surgery was the most interesting from a reconstruction standpoint. The patient was an older male with a progressed facial cancer, which he had delayed seeking treatment for. The cancer progressed to infiltrate most of the structures and tissue within the right half of his face, concentrating mostly under his eye and down his neck. Unfortunately, despite rounds of chemo, surgical removal, and immunotherapy, the cancer was unable to be fully removed. Now the patient was left with a considerable amount of his right face removed and had also had to deal with the devestating effects of facial radiation. The patient had received a skin graft, which had previously failed. It was pretty grusome to see, and what I thought was just facial fat ended up being the patient's exposed skull in some parts of the wound. In this surgery Dr. Spector was evaluating the DermiSphere which he had placed the previous week. DermiSphere is Dr. Spector's own tissue regeneration product. He concluded that the hydrogel was being taken well and added some more. The patient, though, was still left with an open wound at this point. Which had a medicated ointment spread over a piece of gauze to keep it moist. I asked Dr. Spector if he believed that this patient would ever be able to support a skin graft to cover the wound. Dr. Spector answered no and explained this was more of a temporary quality-of-life improvement, as this cancer was going to be terminal. 

With the wide breadth of surgeries I was able to see, I feel like I got an impressive spread of what the OR has available. I really did admire the surgeon's grace. Maybe it would be something I would like to do; I am not sure. I was drawn to science to make things to help people. Medicine is obviously a more tangible and immediate version of that. 

With the rest of the day, I lysed samples for a Western blot, which I will run tomorrow. And I took my day one samples for my ELISA. 

2026.07.16:

Today I set up my Western blot to check my MAVS knockouts again. I also sampled my day two ELISA sample. 

2026.07.17:

Today we imaged the western. Ill make the figure for that over the weekend. I also sampled my day three ELISA sample. Tomorrow I will take day four sample and run the ELISA. As the knocked-out MLKL gene mediated the activation of interferon production, I expect there will be a decrease in interferon shown in the ELISA results. 

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