Single-molecule live-cell imaging

August 28, 2012

Some things in biology can be observed best when concentrating on one molecule and its functions. System biologists will probably not agree and intervene that in biology it’s all about networks and interactions. The presence and concentration of A influences B which increases the concentration of C which consequently down regulates A again. This is all true. However, in single-molecule biology it’s about the functioning and dynamics of (you guessed it) single molecules. When looking at larger systems there is always the danger of missing out elements that might occur only under certain conditions, low concentrations, or that are masked by certain other secondary processes. The weak point of single-molecule studies has always been the fact that complex systems are drastically reduced. Again, you miss-out a lot of stuff even though now you are able to study one molecule in detail.

But: Change will come. As I described in an earlier post super resolution microscopy has been around now for a few years and it is a ready-to-use technique now. For just $ 1,000,000 you can get your own. Theoretically many fascinating research results should have been published  until now. Observe single molecule dynamics in their native environment, what more could you wish for? Indeed some spectacular footage has been produced. Stefan Hell and coworkers, for example, were able to record neurons within the cerebral cortex of a living mouse with a resolution of around 70 nm. Until 20 years ago physics books would have told you that this is impossible. So have a look yourself, right here.

Strangely enough this research at the same time also demonstrates and interesting phenomenon that can be observed when scanning through the live-cell super-resolution microscopy: Most of the time only structurally large (>200 nm) and functionally already known molecules (like neurons) are observed. Further, the temporal resolution is not great and in the order of seconds. Fast moving molecules are still hard to image due to hardware (CCD camera, scanning) limitations. Of course it is very interesting to see how dendrites in the brain expand during learning, but it does not raise any new questions and most importantly does not answer any old ones. I am sure that super-resolution microscopy has a golden  future, but it is important to improve sample preparation techniques, optimize fluorophores even further, and develop sensors that have a shorter integration time for the small amount of photons they are capturing per frame.

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