By Zully Santiago, PRISM Undergraduate Researcher, Spring 2013 through Summer 2014
Part 2: Understand your Experiment Before Beginning
5) Mistake: You quickly get a simple protocol or a recipe for your experiment (or, if you are lucky, you have a kit’s instructions). You immediately perform the experiment, but it fails horribly, and you have no idea what went wrong. You automatically assume the protocol, kit, or recipe is wrong and you did everything correctly.
Solution: Understand the experiment before you do it. Understand what every single chemical does and what role it plays in the experiment. I would say most failures in an experiment are human error by the scientist overlooking something simple but important. My rule, don’t touch a chemical unless you know what it does, how to care for it, what role it plays in your experiment, and how to safely dispose of it. If your experiment uses a kit, thoroughly understand how the kit works. If it is being used for separation, what kind of separation method is it? If it is chromatography, what kind? If it is a gel, how does it separate? What comes out first? Understand all the components of the kit before using the kit.
Often protocols that you find online or in books will not cover important details such as proper care and considerations for the chemicals you are using such as light sensitivity, reactions with moisture and air, temperature concerns, reactions with certain plastics, and so on. Also, keep in mind that time and exposure to the environment can heavily weigh on the experiment. A big overlooked factor is the shelf life of reagents. Remember, often when chemicals are exposed to water, hydrolysis occurs, slowly degrading the chemicals over time. So the chemical may have a long shelf life in its store-bought form, but it may have a very limited shelf life once in solution. This is a big issue for antibiotics and DTT.
Also, repeatedly thawing out and freezing chemicals or proteins can also severely degrade them. Record how often your reagents are thawed out or aliquot small amounts at a time so that you can use those amounts when needed rather than thawing out the whole stock container each time. The protocols and kits usually assume you know what the chemicals do and how to handle them—which often you don’t, so google everything or go to the company website and read up on your experiment and the reagents before handling them. When working with kits, thoroughly read the material that comes with it as it has detailed information on handling all the components of the kit.
6) Mistake: You do an experiment, and it works! Or it fails! Who knows? Either way, you got some type of result…but you didn’t write anything down.
Solution: Again, it’s easy to follow protocols and recipes, but what matters are the specifics! What did you use? What conditions? What temperatures? What amounts? What chemicals? What order? How long? Again, there are so many factors involved in getting results outside of what is mentioned in the protocols. You need to document your steps and what has occurred. Based on your observations you may find better ways of doing the experiment. Or based on your observations, you have resolved a huge problem! However, we won’t know unless you write it down. In my experience, I rarely follow the protocol exactly; often I find better tweaks that provide good yields for my experiment, but I record deviations as well as observations. Moreover, if something goes wrong, often there are good scientific blogs on company websites and third party websites that talk about the same problems you may have had in the lab, so it is important that you record your observations because these forums and blogs may help resolve what went wrong and how to fix it.
7) Mistake: You do the experiment from memory.
Solution: Never do your experiment from memory. You should always have the protocol or your previous observations handy just in case something turns out different. However, the main issue with trying to do an experiment from memory is that you will often forget a step, usually an important one. I see this happen a lot when students make buffers at the total volume desired, but they didn’t adjust the pH, so that added volume was not accounted for.