Reproducibility is key to scientific research. It is, in fact, a hallmark of the Scientific Method, controlling variables until the same result can be achieved time again. There are few things more frustrating than following a simple experimental protocol and not achieving the predicted result. This inevitably leads to wasted time, wasted reagents, and very frustrated researchers.
Nuclear Localisation Signal (NLS) is a peptide sequence that tags certain proteins for nuclear transport. By conjugating NLS to gold nanoparticles (GNPs), this should (in theory) result in the GNPs being transported to the nucleus. This can then be detected using photothermal microscopy. However, actually putting this into practice is slightly harder. Due to the abundance of positively charged residues in the NLS sequence they can cause the nanoparticles to aggregate.
Following the preparation of 10nm GNPs conjugated to CALNN, the next step was to use these for microinjection and then photothermal imaging. For this I used 35mm gridded dishes, this meant that the coordinates of successfully injected cells could be recorded to make the cells easier to find again. The length of time the cells were removed from the incubator had to be kept to a minimum as after extended time periods they would begin to die. After the cells had been injected they were then fixed using paraformaldehyde and stored in the fridge until needed for imaging.