Katie Munechika / Ethel Cesarman - Week 6

This week, I did the immunoprecipitation experiments as planned. Besides NSD2, I also included another histone interactor, STAT3, to further test the capabilities of the IP protocol. Next week, I will be able to perform the western blots to confirm that the IP’s worked properly. The western blots will also help to verify the specificity of the antibodies I used for the IP. For example, I want to make sure that the H1C antibody does not also capture H1E and vice versa. When doing the IP, I was able to confirm that there was an acceptable amount of protein present in the samples after the protein extraction step via a BCA assay, but I was not able to tell if our target histone proteins were included. If the western blot confirms that the IP’s worked properly, then, if time allows, I would like to redo the IP’s with biological replicates, so I can prepare and run them on the mass spectrometer back at my lab in Ithaca.

I continued with mouse work this week as well. Within the past 10 days, some of the wild-type mice underwent bone marrow transplants using bone marrow that we had previously collected from H1 mutated mouse lines. These include knockout of H1C, H1D, or H1E isoforms. Different ratios of the knockout-derived bone marrow were transplanted to test which phenotype would take over. A few days after this process, the mice were immunized to trigger differentiation of B-cells in the spleen. Then, we collected and processed the spleens to separate out and extract the B-cells.

I shadowed Dr. Morales from the Infectious Diseases department again as she saw a few different patients. One new patient she met with had a bacterial lung infection. This was discovered from CT imaging that was done for a different injury prior to this visit. Upon discovery of the infection, a bronchoscopy and lavage performed was performed to collect a sample, which led to the identification of the bacteria as Mycobacterium avium-intracellulare (MAI). This is an extremely common bacteria that usually does not cause any illness, however in this particular case, was the cause of the infection. The patient had an extensive medical history and was also in the process of seeking treatment for liver cancer, so Dr. Morales determined that it was best to consult with the patient’s oncologist and cardiologist before going ahead with any treatment, especially since the patient seemed to be in a stable condition. Generally, this type of infection is treated with three different antibiotics at the same time in case the bacteria becomes resistant to one. One of the medications can cause inflammation in the liver, and the oncologist had suggested radiation therapy for the patient’s liver cancer, so it was best to check with the patient’s other clinicians before deciding on the best treatment and frequent blood work would have to be done to ensure proper liver function. Additionally, Dr. Morales wanted to collect some baseline blood work and sputum samples, as well as eye and ear exams before starting treatment, so that negative side effects of the medications could be detected and managed.