Me pipetting a small quantity of an antibody into a solution to be given to the cells
Over the past two weeks, I have completed my remaining experiments and concluded my internship at the Crawford Lab.
We started the fifth week by expanding our immunofluorescence testing from the cancerous cells to the stromal cell lines comprised of fibroblasts. These cells have a crucial structural and connective role in the body as they are involved in creating supportive frameworks and wound healing. However, these cells can also become an important part of the tumor microenvironment if they morph into a hostile state that encourages the progression and advancement of the tumor.
A picture of my workstation in the Crawford Lab
In these cells, we searched for the same specific protein as we had with the cancerous cells in addition to GM130, a protein which marks the Golgi apparatus, an organelle that functions as the packaging center of the cell and is an important intersection of numerous cellular pathways. The pictures seemed to confirm a binding between the protein and the Golgi across the treatments and in multiple lines.
However, we had noticed in our previous images of the prostate cancer lines that during mitosis, the protein seemed to attach to the microtubules of the mitotic spindle. Furthermore, there seemed to be evidence of the formation of dot-like patches on the spindle fibers, perhaps at the location of their connection to the centromere of the chromosomes.
A dying cell that is in apoptosis
This discovery delighted everyone in the lab, and we decided to continue to focus on this lead. We did so by attempting to draw a connection between the protein (which has been implicated in spindle function) and another that is necessary in the proper attachment of the spindle. To our great joy, we managed to see a colocalization between the two during multiple stages of mitosis; furthermore, there seemed to be differences between the control and fatty acid treatments. This discovery is only in the initial phases, and in the future the lab may well continue to delve into the subject.
A sample from our immunofluorescence. The blue is the nucleus, the red the Golgi apparatus, and the green the target protein
During my final week, we were sadly unable to attempt to overcome the failures in the beginning of my internship experiences to ascertain a Western Blot reading of the cells due to the necessary antibodies being delayed in their shipping. As we did not want to have to use outdated versions, we concluded that I would not be able to do one. However, I was kept busy by starting immunofluorescence testing for my mentor’s project with some of the coverslips I had left over. Furthermore, I was able to sit down with Dr. Crawford and the rest of the team to begin to analyze my findings.
In this time, I also moved in with my final host family, whose father is the son of the initial couple that hosted me. They were also a great group that made me feel extremely welcome. Furthermore, I had the opportunities to visit two of the universities in the area, the University of Chicago and Northwestern; needless to say that they are both amazing institutions. Beyond that, I used the weekends for trips to the beach on Lake Michigan and to continue to visit and explore Chicago, in addition to spending more time with the two Telluride natives in the area and visiting the cinema.
In the end, it was somewhat saddening to leave the lab, but I was also excited to go home and see my family and the dogs. During my last day, it was great as the entire lab went out for a lunch together to say goodbye to me and the other student researchers that had been there with me. Overall, I would like to thank Pinhead, the host families, and especially everyone in the lab that had taken the time to guide and instruct me and had made sure that my time was successful. Thank you for the opportunity.
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