When thinking back on my project, I see it as having three stages. So, I would like to take you along my journey this summer and share some of the ups and the downs that I have encountered along the way.

Stage 1: Cell culturing

Difficulty Level: Easy

My Laidlaw research journey started off with me learning how to culture the osteosarcoma cell line. I had to grow enough cells so as to have enough to carry out all of my experiments. Cell culturing was something I have never done before. It was really intimidating at first. There were so many steps to follow to do even the simplest thing such as feeding the cells, so many protocols and rules, and I was definitely overwhelmed. I was really worried that I would never be able to remember all of the steps and that I would somehow contaminate or kill the entire cell culture that I would spend 2 weeks growing (happy to say that did not happen). But as always, it just takes practice, and it wasn't the end of the world as I first thought it would be. Each day as I was following the protocols, I was becoming more and more familiar with them. At the end of the 2 weeks, my confidence in the lab had grown massively and I felt like I was at the stage where I wasn't going to kill the cells with every step I took. Don't get me wrong, I am by no means an expert at cell culturing and there are A LOT of protocols I need to improve on in the future but I am happy with where I ended up. I grew 8 million cells by the end of those 2 weeks and I didn't contaminate any samples, so yay for me! I then started separating my cells into two groups. Group 1 was the control group, and group 2 was the chemotherapy treated group. 

So lesson number one that I've learnt: Even if something is unfamiliar to you at first, that doesn't mean that it will always stay unfamiliar. Trust in the process and just practice. And if a mistake happens, it doesn't mean that it is the end of the world. The most important thing is that you understood where you went wrong and you learnt from it.

Stage 2: Sample preparation

Difficulty Level: Moderate

The second stage of my project was preparing the samples so they are ready to be put into NMR tubes. This process included: 1) De-plating the cells from cell plates. 2) Removing all of the cell components and proteins, so that just the metabolites are left. 3) Removing as much water from the samples by using a speed-vacuum centrifuge.

As you can tell, there were a lot of things to do to even prepare the samples and it took quite a bit of time. This was mainly due to having to run all 30 of my samples in multiple centrifuges which was very time consuming. This was the point in my research project where I started getting worried because I was already in my 4th week of the research and no where near running the samples in the actual NMR which would take most of my time. I was really struggling with the fact that I wasn't able to follow my original timeline for the project. But in a way, I am happy that this happened. It allowed me to get over my fear of not following my plans to the exact detail and it made me evaluate how I was doing my work. Was I actually spending my time in the lab as efficiently as possible? No, I wasn't. I realised that while I was letting some of my samples run in the centrifuge, instead of waiting I could've been preparing my other samples for the next steps. After starting to implement these changes, it made me realise that I was carrying out my work more efficiently. However, I was still behind schedule and was worried if I'll have enough time to finish everything. Little did I know that my problems would get even worse after this...

So lesson number two: If something isn't working the way that it's supposed to, take a step back. Breathe. Think. Don't let the panic consume you. There is always a solution to things. Evaluate how your work is going, and adapt to the situation.

Stage 3: NMR

Difficulty Level: High

After making all of my samples, it was time to finally run them in the NMR spectrometer. The first week was me learning from my supervisor how to run the machine and optimise all of the conditions for running my samples. It was really fun learning everything as it was something I have never done before. But it was definitely intimidating as the machine was really expensive and I didn't want to do something and make it blow up. Happy to report I have not caused an explosion yet! Everything was going well up until now. I was becoming more comfortable with running my samples, but then the biggest hurdle came. The quality of my scans were not very good. The reason for this was that room where the machine was located was experiencing technical difficulties and this was negatively affecting the quality of my results. At that point, I felt so defeated. I was running really behind my original schedule, I was stressed that I wouldn't get everything done in time and now the quality of my scans was not good either and the most frustrating part was that I couldn't do anything about the quality of my scans. Everything was just going wrong. I felt like my entire project was a failure. But even from all of it going wrong, I've learnt a very valuable lesson.

So finally, lesson number three: If everything is going wrong, and you can't do anything about it, don't let that dishearten you. Do the most that you can, and at the end of it you'll know that you've given your all. And that is enough. It is enough that you've done your best despite all the challenges, and be proud of yourself for not giving up.

Tea Stapar, 2023 Laidlaw Cohort, Biochemistry student