I had a meeting today with Prof. Klaas Hellingwerf and Jos Arents at he University of Amsterdam Swammerdam Institute of Life Sciences. They work with differently colored fluorescent proteins and bacteria as well, so they actually have all the expertise I could ever need to this project when it comes to biotechnology. We’ve discussed possible issues with the pH-dependent fluorescence change, and also new exciting approaches on how to make this fluorescence change happen.
First we’ve touched the issue of UV light being harmful for cells, and thus especially for single-cell organisms such as bacteria, but also potentially to visitors of the installation. But, UV light is needed to excite the fluorescent protein, so as to emit fluorescent light. Fortunately, the harmful frequency light range and the exciter light range doesn’t overlap totally. Thus it’s possible to chose a light, closer to the visible spectrum, that is not as harmful but still excites the fluorescent stuff.
One of the alarming remarks was that in the Wilks article, it is shown that a change of pH between 5.5 and 7 will cause a three-fold change in the level of fluorescence for the fluorescent protein. Unfortunately, this level of change is probably hard to detect by the human eye. As the bacteria itself is already growing, I’ll be able to see if the change is big enough to see by the human eye.
So far, I’ve been thinking on dropping acid and other chemicals on the bacteria to change the surrounding pH value. But during the meeting an alternative approach was suggested – to have a photo-sensitive intra-cellular pH value change. That is, one could make the bacteria change its internal pH value, and thus its level of fluorescence by shining light on it. This would make the installation so much simpler and nicer. And would also open the door for the bacteria-to-go aspect.
An additional idea that came up was to use Bacillus Subtilis instead of E. Coli, which has a protein called YtvA. This protein is in fact a receptor for blue light, and could trigger expression of GFP as a response. Thus again, if one shines light on the bacteria, it would start to become fluorescent in about 30 minutes. As this GFP is apparently quite unstable, the fluorescence would fade away quite fast – resetting the bacteria into its initial state.
We’ve also touched the issue of sustainability for such an installation, and unfortunately it doesn’t particularly look good. Bacteria on a solid plate are active for a short amount of time only, like a day, after that, they turn into a stationary state. In the latter state, they don’t really express DNA any longer, and everything is a lot slower. One could put them into a liquid base, so as to replenish the food supply, etc. But then the liquid has to be stirred constantly, so that there’s enough oxygen & stuff all around – which in fact creates a homogeneous environment. This is contrary to what we want to do to create a display: have different spots look differently.
But all in all, this was a very fruitful meeting. I’ll know if the level of fluorescence change as a response to pH change is human-visible in a few days, and then we can go forward…