Vivian Sun :: Week 6 :: The Grand Finale
Week 6 was my final week! The time went by faster than I ever could have imagined - six weeks seemed a daunting stretch of time at first, but I realized how short it really was as I started my sixth week at JHU.
I wrote up and finished my bone marrow monocyte isolation protocol over the weekend and began my final week by going over the protocol with Snow. I got a clearer sense of what we would be doing, why we were doing each step, and what the overall purpose of the experiment; she answered all of my lingering questions and provided helpful tips for me to keep in mind in anticipation of and in preparation for the experiment.
For clarification purposes, my new protocol was called "Bone marrow monocyte enrichment using the Miltenyi bone marrow monocyte isolation kit" (Miltenyi is the name of the company that makes the enrichment/isolation kits). We wanted to look at how many monocytes could be obtained from the bone marrow, as well as test out the aforementioned Miltenyi kit to see how effective it would be at maximizing the monocyte yield. The experiment would take place on Thursday.
I made up the MACS buffer, found and laid out the syringes and syringe tips, cleaned up and obtained the sacrifice table and instruments, and set up the well plates and 1XHBSS. I did all that I could prior to the experiment to make Thursday's sacrifice, harvest, and processing go as smoothly as possible and as efficiently as possible.
Ms. Cozine visited me on Tuesday; it was a pleasant break from the presentation and experiment preparation! I loved having Ms. Cozine as an AP Chemistry teacher last year, so I was excited for her to come down to Baltimore and visit me. I had a rough presentation put together, so explaining my summer project was actually great presentational practice for me, to boot!
In addition to preparing for the experiment, I spent a large portion of my time working on my presentation and EXP poster. The lab meeting on Friday would be for "Summer Student Presentations" - aka Katie and myself presenting on the work we had done during our time at the lab. I would be presenting on both of my experiments, and Katie would be presenting on the experiments she conducted/helped out other lab members with.
In terms of creating the presentation, I used Prism (an alternative to Excel that is much better at creating graphical representations of data) to display and make the pie charts and bar graphs I would be showing. The presentation took a long time to finish and polish - I wanted to display the data as clearly as possible with minimal words on each slide.
As I worked on my presentation and data analysis/visualization, Snow was figuring out t-SNE, a cutting-edge technology that was a plugin for FlowJo (the software used to analyze the flow cytometry data). Each time she wanted to compress the data and create a final graph, the t-SNE process lasted hours (often overnight), but the end result was gorgeous. t-SNE is basically a graph in which the multiple dimensions of the data set are compressed into two dimensions.
Thursday rolled around and it was a very long day - I was in the lab for about 12 hours. I started off by sacrificing the mouse and harvesting its spleen and bone marrow. Snow and I then processed the spleen and bone marrow cells and ran magnetically-labelled cells through the LS magnetic selection column. We stained compensation and isotype controls, as well as the samples, and we went to do flow cytometry around 3 in the afternoon. It was my third time going to the tenth floor to run flow cytometry. I had always watched in the past, maybe clicked a few buttons here and there, but this time it was just Snow and me - and I got to run a majority of the flow cytometry! I learned how to not only run the machine but also how to set up and run the software for data collection.
After obtaining the raw data, I analyzed it with FlowJo, Excel, and Prism. It was 5 pm, so I didn't have a huge amount of time for data analysis or creating graphs/visuals, but I did what I could on PowerPoint for Friday's meeting/presentation.
Snow took the bone marrow data and created a t-SNE graph with it. It took 5-6 hours since it was a smaller data set than the one she had tried out before, but the graph she created was gorgeous! In this case, the t-SNE graph is showing the different cell type populations if the cells therein overlap. It's a visual representation of the cell types in the bone marrow, which I think is extremely cool!!
All the purple areas are the Ly6Clo monocytes, aka the monocytes that patrol the blood streams. The yellow areas are the Ly6Chi monocytes, aka the pro-inflammatory monocytes that have a short lifespan. CXCR4 is a chemokine receptor that, when upregulated in cells in the bone marrow, leads to retention of these cells in the bone marrow (green). CCR2 is another chemokine receptor that, when upregulated, leads to cells egressing from the bone marrow (red). You can see that the yellow area also has some green overlap, so the Ly6Chi (yellow) cells also may be CXCR4+, meaning that the cells would stay in the bone marrow. There's also some red overlap with the yellow, meaning that some yellow Ly6Chi may be CCR2+ and would likely leave the bone marrow.
On Friday, Daniela (my PI) and Katie both brought cakes, which was a welcome surprise! Cake is always wonderful. My presentation went better than expected; I also received some pointers for what to include in the PowerPoint for when I present it at school. Some suggestions were also about clarity and making sure that the material presented is concise, but overall, I'd call my presentation a success! After the meeting, we went outside to take a lab group photo (or 10).
I took some final photos with Snow and Daniela. I said my goodbyes to everyone at the end of the day on Friday and went to turn in my volunteer badge to the Volunteer Services JHU hospital office. My six long but exciting and educational weeks of lab were done! I did get an invitation to come back next summer, and I definitely would love to!
I wrote up and finished my bone marrow monocyte isolation protocol over the weekend and began my final week by going over the protocol with Snow. I got a clearer sense of what we would be doing, why we were doing each step, and what the overall purpose of the experiment; she answered all of my lingering questions and provided helpful tips for me to keep in mind in anticipation of and in preparation for the experiment.
For clarification purposes, my new protocol was called "Bone marrow monocyte enrichment using the Miltenyi bone marrow monocyte isolation kit" (Miltenyi is the name of the company that makes the enrichment/isolation kits). We wanted to look at how many monocytes could be obtained from the bone marrow, as well as test out the aforementioned Miltenyi kit to see how effective it would be at maximizing the monocyte yield. The experiment would take place on Thursday.
I made up the MACS buffer, found and laid out the syringes and syringe tips, cleaned up and obtained the sacrifice table and instruments, and set up the well plates and 1XHBSS. I did all that I could prior to the experiment to make Thursday's sacrifice, harvest, and processing go as smoothly as possible and as efficiently as possible.
Ms. Cozine visited me on Tuesday; it was a pleasant break from the presentation and experiment preparation! I loved having Ms. Cozine as an AP Chemistry teacher last year, so I was excited for her to come down to Baltimore and visit me. I had a rough presentation put together, so explaining my summer project was actually great presentational practice for me, to boot!
 |
| from left to right: Snow, myself, Ms. Cozine |
In addition to preparing for the experiment, I spent a large portion of my time working on my presentation and EXP poster. The lab meeting on Friday would be for "Summer Student Presentations" - aka Katie and myself presenting on the work we had done during our time at the lab. I would be presenting on both of my experiments, and Katie would be presenting on the experiments she conducted/helped out other lab members with.
In terms of creating the presentation, I used Prism (an alternative to Excel that is much better at creating graphical representations of data) to display and make the pie charts and bar graphs I would be showing. The presentation took a long time to finish and polish - I wanted to display the data as clearly as possible with minimal words on each slide.
As I worked on my presentation and data analysis/visualization, Snow was figuring out t-SNE, a cutting-edge technology that was a plugin for FlowJo (the software used to analyze the flow cytometry data). Each time she wanted to compress the data and create a final graph, the t-SNE process lasted hours (often overnight), but the end result was gorgeous. t-SNE is basically a graph in which the multiple dimensions of the data set are compressed into two dimensions.
Thursday rolled around and it was a very long day - I was in the lab for about 12 hours. I started off by sacrificing the mouse and harvesting its spleen and bone marrow. Snow and I then processed the spleen and bone marrow cells and ran magnetically-labelled cells through the LS magnetic selection column. We stained compensation and isotype controls, as well as the samples, and we went to do flow cytometry around 3 in the afternoon. It was my third time going to the tenth floor to run flow cytometry. I had always watched in the past, maybe clicked a few buttons here and there, but this time it was just Snow and me - and I got to run a majority of the flow cytometry! I learned how to not only run the machine but also how to set up and run the software for data collection.
After obtaining the raw data, I analyzed it with FlowJo, Excel, and Prism. It was 5 pm, so I didn't have a huge amount of time for data analysis or creating graphs/visuals, but I did what I could on PowerPoint for Friday's meeting/presentation.
Snow took the bone marrow data and created a t-SNE graph with it. It took 5-6 hours since it was a smaller data set than the one she had tried out before, but the graph she created was gorgeous! In this case, the t-SNE graph is showing the different cell type populations if the cells therein overlap. It's a visual representation of the cell types in the bone marrow, which I think is extremely cool!!
 |
| t-SNE depiction of our data (legend below) |
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| legend of the t-SNE graph above. |
On Friday, Daniela (my PI) and Katie both brought cakes, which was a welcome surprise! Cake is always wonderful. My presentation went better than expected; I also received some pointers for what to include in the PowerPoint for when I present it at school. Some suggestions were also about clarity and making sure that the material presented is concise, but overall, I'd call my presentation a success! After the meeting, we went outside to take a lab group photo (or 10).
 |
| We did have a more serious standing group photo, but I liked this fun selfie of the lab more! Daniela (my PI) is the one taking the selfie. From left to right, behind Daniela: Katie, William (in the blue), Taejoon (in the white), Paul (in the purple), Snow (in the beige), and myself (in the white/blue). |
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