Steve Mirsky: Welcome to this third episode of our special Nobel Prize editions of Science Talk, the podcast of Scientific American. I am Steve Mirsky. Today, the Chemistry Prize.
Staffan Normark: This year's prize is about how the optical microscope became a nanoscope.
Steve Mirsky: Royal Swedish Academy of Sciences Permanent Secretary Staffan Normark.
Staffan Normark: The Royal Swedish Academy of Sciences has decided to award the 2014 Nobel Prize in Chemistry to Dr. Eric Betzig at Howard Hughes Medical Institute, Aspen, USA; Professor Stefan Hell at Max-Planck Institute for Biophysical Chemistry, Gottingen, and the German Cancer Research Center, Heidelberg, Germany; and Professor William Moerner at Stanford University, Stanford, USA for the development of super-resolved fluorescence microscopy.
Steve Mirsky: A little background as you continue to listen, you'll hear the members of the committee refer to the Abbe diffraction limit or the Abbe condition. German physicist Ernst Abbe determined that you can only look at an object in a microscope in focus up to a certain size, related to the wavelength of the light that you're shining on the object. The new Nobel laureates have come up with ways to get around that limit. A key way is that they're not really looking directly at the object; to the tiny thing that they do want to look at they add numerous little tags that fluoresce, that glow, then they observe that fluorescence. If you add enough little tags you see finely-focused details, just as if we dressed you in a suit covered with tiny little light bulbs, maybe the blue diodes, whose inventors won yesterday's physics prize, and then watched you walk around in a dark room, we could see exactly where you are, how you're moving your limbs, et cetera.
Okay, back to the announcement.
Staffan Normark:Professor Sven Lidin will now give us a short summary. Please.
Sven Lidin: Thank you, Staffan. Could someone please do something about the resolution? That's a question that microscopists have asked for many years. Ever since the invention of the microscope this has been the challenge. We see new things, but we see that there are more things to see.
In 1873 Ernst Abbe proved that the theoretical limit to a resolution is about half the wavelength of the light used. For visible light that translates to about 200 nanometers. Now a single human hair is about 100 micrometers. That's 500 times larger than this limit. So objects of that size are very easily studied with conventional microscopy. But most of the processes in chemistry and biochemistry take place at length scales that are much smaller. A typical bacterium is about 200 nanometers across, corresponding to the Abbe limit, which means that if we study bacteria in the light microscope they will be featureless blobs.
This prize celebrates the circumvention of the Abbe condition. And three scientists have contributed to break this limit in two different ways: Stefan Hell developed stimulated emission depletion microscopy, STED; and Eric Betzig and W.E. Moerner laid the foundations for a single fluorophore microscopy. Now why is this important? Electron microscopy has a much higher resolution and can be used to study structures in atomic resolution, but electron microscopy does not allow us to study living cells or the processes within them. The work of the laureates has made it possible to study molecular processes in real time. It's been used to study the dynamics of transcription and translation, that is the reading of DNA and the transference from that to proteins. It's enabled us to look at how proteins associated with a disease aggregate, for example, in Alzheimer's, Huntington's, and Parkinson's disease. And it's even shown us the structural dynamic changes to neurons in the brain that takes place during learning processes.
Super-resolution microscopy doesn't only tell us where, but also when and how, and that is the greatness of this development. Biology has turned to chemistry, chemistry has turned into biology. Guesswork has turned into hard facts, and obscurity has turned into clarity. Now we can observe E. coli in all the glory of super-resolution without having to kill them, slice them, fixate them, and subject them to intense radiation and high vacuum. They can be studied in real time, while they live long and prosper.
Staffan Normark: We're now trying to get hold of Professor Stefan Hell, and I think we have him on line from a phone in Germany. Good day, Professor Hell. Congratulations.
Stefan Hell: ______ ______ ______. Thank you. Thank you so much.
Staffan Normark: So, Professor Hell, I'm sitting here in the session hall at the Royal Swedish Academy of Sciences, and we have a number of journalists from all over the world that are here, and I think some of them are eager to ask some questions.
Stefan Hell: Sure.
Staffan Normark: And here comes the first one.
SVT: Hello, Professor Hell. [Speaking foreign language]. Congratulations.
Stefan Hell: Danke schön. Thank you.
SVT: This is Swedish Television, SVT, your live – our live broadcast. Could you – something has already been said about your work. Could you describe the significance of it in your own words, please?
Stefan Hell: Yes. I mean light microscopy is very important to the life sciences because the use of focused light is the only way that allows you to see living things, however, the resolution of light microscopy was fundamentally limited to about a fifth of 1,000 nanometer, and you want to see – they brought in distributions at much higher resolutions, at much smaller scales, which was simply impossible and definitely not in live cells. Now what we have found is that you can overcome this limit, you can see details at much, much higher space resolution, and this of course discloses how the cell works at the nanometer scale, so that's at the molecular scale, at the macromolecular scale, and is of course very, very important to, (a) understanding how the cell works, and (b) understanding what goes wrong if the cell is somehow diseased, if something, you know, diseased that thing or something. So it's very important for understanding the physiology at the cellular and of course understanding disease at the cellular level.
SVT: If I may continue, could you say something about how you got the idea?
Stefan Hell: Yeah. So actually the reason why I came across this diffraction problem was that I worked with my PhD thesis on light microscopy - ____ ________ microscopy, but not for the life sciences, not for fluorescence. It was basically for microlithography inspection. And I got bored with the topic and I found this is kind of 19th century physics and I was wondering whether there is still something profound that could be made with light microscopy. And so I saw that diffraction barrier; that's the only say important problem that has been left over that people have saw that it would not be beatable, you cannot do anything about it, you have to resort to electron microscopy if you want to see the nano scale.
And so I got attracted to this project and I started thinking about well there's really no, say, physics or physics in chemistry or chemistry to sort out the problem. And eventually I realized that there must be a way by playing with the molecules. Trying to turn the molecules on and off allows you to see adjacent things that you couldn't see before.
Staffan Normark: Okay, thank you. Do we have some other questions to Professor Hell?
Joanna Rose: Hello, and congratulations, Professor Hell.
Stefan Hell: Thank you.
Joanna Rose: My name is Joanna Rose. I am from the Swedish popular science magazine Forskning & Framsteg.
Stefan Hell: Right.
Joanna Rose: And how big was the challenge to put the molecules on and off?
Stefan Hell: Well, it is, of course, something that you have to realize first, you have to realize that there are methods of turning molecules on and off. Now I realized, and this was explained by Professor Mäns Ehrenberg, that you can turn molecules off by a phenomenon called "stimulated emission," that's a very, very basic phenomenon; every student learns about that in the first year. And after I realized that you can turn it on and off with stimulated emission I knew that there were also other mechanisms for turning molecules on and off that are ______ described in the literature, for example pumping things along with doxates versus trantizimation and so on. And so there are many, many ways of playing the same game, and this turned out to be, of course, very, very successful. So it's now a whole field and you can play this concept in many, many ways.
Joanna Rose: Can I continue? Just you mentioned actually in our magazine for about two years ago that-
Stefan Hell: Yes, I remember.
Joanna Rose: -you were really on the verge of giving up.
Stefan Hell: That's right. I mean it's certain to say that the scientific community wasn't very receptive to the idea of overcoming the diffraction barrier because people believed the barrier has been around since 1873 and the resolution is what it is and doing something about it is, pardon me, kind of crazy and not very realistic to do. But having realized that you do not overcome the diffraction barrier just by trying to change the waves of light, but by playing with the molecules, changing the states of the molecules. I knew that this was going to work, because I couldn't find a serious, say, physical or physically chemistry a reason why it would not work out. So this is why I was so confident and kept on going, despite all the issues and problems that came up in the interim.
Staffan Normark: Okay, thank you. Do we have another question as well? Over there.
China Radio: ______ _______, freelance for China Radio and also Green Post. And I thank you- congratulations.
Stefan Hell: Thank you.
China Radio: And you sound still very calm actually. So did you expect you'll win the prize? How did you feel when you hear the news?
Stefan Hell: No, I was totally surprised. I was totally surprised; I couldn't believe it. But fortunately I remembered the voice of Professor Normark, and so I realized that this is real. But it took me a while to realize it, I must say.
Staffan Normark: Okay, thank you very much, Professor Hell. And we are looking forward to meet again in December for the Nobel Prize ceremony. And our warmest congratulations.
Stefan Hell: Thank you.
Staffan Normark: So now I turn to you, or do you have any other questions or further questions to our experts?
Female: [Speaks in foreign language]
Mäns Ehrenberg: Can you take the answer in English?
Steve Mirsky: This question is fielded by Nobel chemistry committee member Mäns Ehrenberg.
Mäns Ehrenberg: And yeah, the question was is there any limit that is still valued for the resolution you can get. And the answer is essentially there is no strict physical limit left. However, if you want to have a very high-resolution image of an organelle by fluorescence labels you must have labeled it properly. So you have labels everywhere essentially. And so there may be a practical limit, but it's much, much lower than the limit that was set by Abbe or understood by Abbe in 1873.
Mäns Ehrenberg: Six, 1876. Today it was described. Yeah?
Female: What is that _______?
Mäns Ehrenberg: And it was so well-believed, this limit. This is why Stefan Hell is – you know, I almost gave up, because he was up against a physical limit, and most people who are up against a physical limit do not survive professionally.
Steve Mirsky: After the announcements Swedish science journalist Joanna Rose spoke with Sven Lidin. Some people joked already that this chemistry prize was for a physics invention that's useful in biology. Lidin will explain that though there's some truth in that idea, the chemistry aspect of super-resolved fluorescence microscopy is a really big deal.
Joanna Rose: Sven Lidin, you are the German of the Nobel committee in chemistry that just awarded three Nobel laureates who have developed a new super-resolution microscope. Can you tell us what this super-resolution is, how big or how small is it?
Sven Lidin: Well, the super-resolution refers to Abbe's limit, which states that a microscope can never be better, give better resolution than half the wavelength of the light used. And the methods that has been developed by this year's laureates shows that this limit is not strictly enforced. Actually it can be not only broken, but superseded to any resolution. It's not applicable to any system. It means that we need to do some alterations to the system, first of all we need to make them fluorescent. But if that is possible there is – the physical limit is much, much further down, which means we can study much, much smaller objects. And this turns microscopy into a chemical technique, while it used to be a biological technique.
Joanna Rose: So what new worlds can you see with this new microscope?
Sven Lidin: Well, I think the easiest way to look at it is to think back to when microscopes were first used. So the first images that were drawn from microscopes, that started microbiology. Now what is happening now is that because we can see individual micromolecules moving about in a living cell, we can study chemistry at a single molecule level and in real life. And this is very, very important to chemistry, because chemistry has traditionally been about studying a large number of molecules and the effect that they have. Here we can look at a single molecule as it is active in a chemical system. That means that rare events can be studied in a very different way, reactions can be studied as they happen, not as the end result, but actually as they take place. It opens entirely new possibilities for chemistry and for biochemistry.
Joanna Rose: So this is like you previously could see an anthill and now you can follow every single ant.
Sven Lidin: You can look at the legs of the ants and you can look at the damage done to the legs of the ant. Yes. And it's a fascinating subject, I think, because it is really a prize that goes into all the prize areas of the scientific Nobel prizes. It has great potential in medicine, it is a prize which has a lot to do with physics, but it's also a prize that has a lot to do with chemistry. It's one of these prizes that eradicates the borders between the subjects.
Joanna Rose: When you think about microscopes today there are already microscopes with this super-resolution, and it's different kinds of electron microscopes. So what is the difference?
Sven Lidin: The big difference is that a light microscope is much less damaging to what it studies. If you want to study a cell by electron microscopy, first of all you have to slice it very thinly, because electrons only penetrate through a very small amount of matter. That means the cell is dead and you have to slice it, you have to fixate it with various kinds of stains to make the parts within the cell visible. This means that you do not have the dynamics, you cannot study real-life processes. And we have learned a lot from electron microscopy, it's a fantastic technique, but this takes us into the dynamic realm, and that is where real chemistry happens.
Joanna Rose: You don’t want the dead ants, so to say, to study.
Sven Lidin: Dead ants are also interesting and we learn a lot from dead ants, but live ants are better, and it's better for the ants too.
Joanna Rose: This prize is awarded actually for two different microscopes. Do they see different things as well?
Sven Lidin: It's really two different techniques and they can be used on the same systems, but they also have their own limitations. You asked before about the fact that what are the new limitations; well, for anything that works with fluorescence, which both methods do, you need to label the target of your study densely with fluorophores, and then you can see every individual fluorophore, and thus the shape of the object that you are studying. Now the kind of fluorophores that you can use are different for these two studies. With the STED method that was developed by Stefan Hell you need to quench the fluorescence using a very powerful laser that takes away most of the fluorescence. This can cause damage to living cells, simply photon damage, the same that you will encounter if you go to the beach without enough sun protection factor.
Joanna Rose: Too much light.
Sven Lidin: Too much light is not good for you. So there are certainly limitations to both techniques, and they are slightly different, and therefore there is a complementarity here, but they also work very much on the same kinds of systems.
Joanna Rose: And there is this common picture of inventors like artists, mostly poor and hungry and passionate. How do the – this year's Nobel Prize laureates fit into this picture?
Sven Lidin: Well, I think they fit very well. I think it fits particularly well when these inventors are doing something that is common knowledge is that it doesn't work. And so it's an uphill struggle until they succeed. This is quite common actually in science, and I don’t think it's a bad sign. Science is rather protective of its paradigms, and it needs to be protective of its paradigms. Now our laureates this year have changed those paradigms, and that is good. But at the same time that these people have worked, we have hundreds of people who have worked on trying to change paradigms that stood up to the challenge, the paradigms stood up to the challenge. And it is important to defend these paradigms as well. These are what builds science. Every now and then we need to change them, but if we change everything at the same time then the ground on which we stand starts rocking. And most of these things are actually true. Most of the paradigms have a very long lifetime because they describe nature in a correct fashion. In this case the other condition is still valid. These persons simply found a way around them. That's very good.
Joanna Rose: So you've got to be quite the stubborn scientist-
Sven Lidin: You have to be stubborn. You have to have a very high opinion of your own ideas. And you have to have stamina. Those are very important character traits when you're battling with giants.
Joanna Rose: But did they give up? Do you know that?
Sven Lidin: There are stories from the autobiographies of both Eric Betzig and Stefan Hell that there were certainly periods of doubt. Eric Betzig left academia for some time because he considered that he was at a dead end. And Stefan Hell moved around a bit in order to find a place where he could conduct his work, and he needed to get a few theoretical studies in place to show that his ideas were valid.
So yes, I think they were both on the verge of giving up, but they came back to this question because it was so interesting, it was so alluring, and the possibilities were so fascinating.
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