Team:UiOslo Norway/Discussion
Discussion
Sadly we were not able to prove the concept of a biolaser with our gain medium. We tried to use pure sfGFP in a sample as it had a more concentrated solvent of sfGFP to prove lasing, but was not successful. As well we did not get an result for the yeast cells.
We did see some light being emitted with the corresponding wavelength for sfGFP, with the sfGFP solvent, but it was not strong enough for us to oberve lasing within the gain medium.
XX ref to single cell article that manage to have lasing within a pure solvent of GFP? XX
When trying to figure out what went wrong, we quickly looked at our mirrors. Due to lacking of equipment with the correct specs at UiO, we joyfully got some mirrors sponsored by Thorlabs. Due to the fast approaching deadline we had to improvise by using filters and concave mirrors to get the “Shortpass Dichroic Mirrors”-effect. This made us change our initial plan for the set up, and we knew it would be hard to get the mirrors aligned correctly. As we also did not have a optical bench we made due with what we had, but in the end it proved not successful.
Another reason might be that we did not manage to get high enough intensity from the LED. Still by trying to concentrate (and saving) the light, and by adding another LED, we did not see any trace of lasing with the CCD camera. We did see some light in our spectrometer, but we did not manage to catch the light from the CCD camera and therefore it’s difficult to say what kind of properties the light had.
Lastly, when we finally felt confident in our set up, the sfGFP was getting old and it actually emitted some red light it not did before. At this point we tried once more with the yeast but was still not able to catch anything with our CCD camera and there was no time making any new sfGFP.
Other mentions: why red light, why not able to measure the light when we could see it with our eyes?
We were able to properly clone and submit the composite part, as well as the NMT1-promoter we made, but due to time constraint we were unable to test our system in yeast cells. Because of this, we used an existing expressions system to grow GFP-expressing yeast cells and use that to test our laser. This, combined with the sfGFP-solutions, gave us the material we needed to be able to conduct proper experiments with our setup.
We did see some light being emitted with the corresponding wavelength for sfGFP, with the sfGFP solvent, but it was not strong enough for us to oberve lasing within the gain medium.
XX ref to single cell article that manage to have lasing within a pure solvent of GFP? XX
When trying to figure out what went wrong, we quickly looked at our mirrors. Due to lacking of equipment with the correct specs at UiO, we joyfully got some mirrors sponsored by Thorlabs. Due to the fast approaching deadline we had to improvise by using filters and concave mirrors to get the “Shortpass Dichroic Mirrors”-effect. This made us change our initial plan for the set up, and we knew it would be hard to get the mirrors aligned correctly. As we also did not have a optical bench we made due with what we had, but in the end it proved not successful.
Another reason might be that we did not manage to get high enough intensity from the LED. Still by trying to concentrate (and saving) the light, and by adding another LED, we did not see any trace of lasing with the CCD camera. We did see some light in our spectrometer, but we did not manage to catch the light from the CCD camera and therefore it’s difficult to say what kind of properties the light had.
Lastly, when we finally felt confident in our set up, the sfGFP was getting old and it actually emitted some red light it not did before. At this point we tried once more with the yeast but was still not able to catch anything with our CCD camera and there was no time making any new sfGFP.
Other mentions: why red light, why not able to measure the light when we could see it with our eyes?
Biological system
The extracted sfGFP had impurities as we can see in the results of SDS-PAGE for the first experiment. Therefore, not all 35.35 mg is sfGFP, but for our purposes, it was good enough. The main goal is to use yeast cells that are expressing sfGFP as a gain medium, and in that case as well there are impurities in the form of subcellular components of yeast cells. Therefore, that should not be a problem for our purposes.We were able to properly clone and submit the composite part, as well as the NMT1-promoter we made, but due to time constraint we were unable to test our system in yeast cells. Because of this, we used an existing expressions system to grow GFP-expressing yeast cells and use that to test our laser. This, combined with the sfGFP-solutions, gave us the material we needed to be able to conduct proper experiments with our setup.
