If someone had told me eight weeks ago how wrong my expectations about my project were going to be, I would have laughed or cried. I would get by on my previous pre-reading. Wrong. I was going to walk into a lab with a pre-prepared experimental procedure in place. Nope. I was going to stick exactly to my schedule throughout the six weeks. Laughably incorrect. After a few days of training, I was going to be independent in the lab. Woefully inaccurate.
I had read up on astrocytes and senescence when I first created my project proposal. I naively believed that this limited background reading would carry me through. I knew that having knowledge of the field I was walking into would be important but didn’t realise how difficult and challenging it would be to ground myself in astrocyte senescence and Alzheimer’s. I quickly realised that I had only scratched the surface of a complex and nuanced area of research in my preliminary reading. I spent the first week toiling away on Google scholar and PubMed, trying to gather and read as many papers relating to this topic as possible. I spent hours running through what seemed like an endless list of papers. In that first week I also thought that once I started lab work, all the reading would be behind me. Again I was wrong, my reading continued throughout the majority of my weeks in the lab. As with a lot of experiments, I had a lot of downtime between each stage. I found myself trying to gather as much information as I could during these gaps. Background reading is not just something that is completed at the start of the project but is an ongoing process as and when you need to gain that added depth needed to your understanding.
There is a lot that can go wrong in immunohistochemical experiments (many of which I am now intimately familiar with) but within these areas of error there is a lot of modularity in regards to experimental design. If a tissue isn’t showing the correct staining that you expected, it could be due to a number of possible factors, including: incorrect antigen retrieval, the tissue could have been treated too harshly which would degrade the proteins you’re looking for or not enough which wouldn’t free the protein from the bound state it is stored in; incorrect blocking, the protein of interest could be blocked or background proteins may be improperly blocked. You get the idea. Due to this modularity, optimisation was quite difficult. I began tweaking different parts of the experiment to continue to improve the visibility of the proteins I was looking for. I quickly realised how long optimisation takes. I could only change one variable at a time and it took two days to see the result of that change.
Senescent Astrocytes stained using Beta-Galctosidase Immunohistochemistry.
What I thought would take two weeks soon began eating into my time for another type of experiment I had planned. I learned something that I think a lot of people learn from this process, things do not go to plan. It is impossible to predict every obstacle that will arise throughout a project and properly allot time for any action needed to solve this problem. That being said, I was way too optimistic in the planning stage and at the beginning of my project in the amount of work I could complete in six weeks. What was supposed to be two weeks of one experiment type, two weeks of another experiment type and two weeks of analysis quickly turned into a week of solid reading, five weeks of optimisation of an immunohistochemistry protocol and two additional weeks of actually completing one of the experiments I had set out to do. Biological, and scientific research in general, is a slow process and it is unrealistic to have a world-changing, tangible output at the end of six weeks.
People say there are no such things as stupid questions but I feel as though I spent my time in the lab asking pointless and obvious questions. I didn’t realise at the time, just how reliant I would be on Hugh, my lab supervisor. I thought that I would get a week of training, get a feel for the experimental procedure, then potter away by myself for the rest of my time in the lab. I was dependent on Hugh for the first month of my project and when I attempted my first experiment completely independently after four weeks it still felt daunting. I have realised just how collaborative scientific research actually is, even in neuroscience which can sometimes feel like a disjointed field. I won’t always be relying on people to show techniques or where to find something in the lab but science is a synergetic process and the last eight weeks have cemented just how essential team-work and collaboration are.
The dreaded 24-well plated with mouse brain tissue sections.
The last eight weeks of my summer have been challenging but extremely rewarding. I have learned many new lab techniques, gotten a feel for researcher life and made my first steps on my leadership journey. Thanks to Dr. Colm Cunningham for the opportunity and support. Thanks to everyone in the Cunningham lab, especially Hugh, for all their help throughout this project.