MTT Assays

Before I get into the project about mitotic bookmarking, I’m getting to a good stopping point in my main project. Right now, I’m doing a simple experiment called an MTT assay. MTT is a type of yellow salt that we can use to determine the activity of cells. In healthy cells, mitochondria inside the cell metabolize the MTT into a molecule called formazan, which is purple. We can then measure the light emitted by the sample to determine what concentration of formazan exists in the sample. If we observe more of a purple color, more healthy cells exist in our sample; the less purple it is, the fewer healthy cells there are. I’ve been using this type of experiment a lot to test different combinations of drugs. If we don’t observe much of a purple color, we know that our drugs are effective in killing cells. Right now, I’m using the same concentrations of drugs as I have before but I’m modifying the cells a little bit by deleting specific genes. To do this, we are using something called small interfering RNA (siRNA). When siRNA is released into a cell, they bind to messenger RNA, which are the intermediates in the process of making proteins from DNA. As a result, the mRNA is degraded so that no proteins are able to form, thus leading to a variety of potential cellular effects. This sounds difficult to achieve experimentally but it’s not that hard. You just mix a few solutions of reactants. It’s one of the most common things we do. 

The bad thing about doing MTT assays is that they are tedious. It involves a lot of pipetting of small volumes of solution. It can take an hour or two to fully set up a 96 well plate and add drugs to the cells. It’s kind of annoying but it’s one of the simplest experiments I’ve done. And it usually gives pretty good results, which is nice.