Microscope work wins Nobel Prize in Chemistry

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Microscope work wins Nobel Prize in Chemistry

#1  Postby DougC » Oct 08, 2014 9:59 pm

B.B.C. Article
The 2014 Nobel Prize in Chemistry has been awarded to a trio of researchers for improving the resolution of optical microscopes.
Eric Betzig, Stefan Hell and William Moerner used fluorescence to extend the limits of the light microscope.
The winners will share prize money of eight million kronor (£0.7m).
They were named at a press conference in Sweden, and join a prestigious list of 105 other Chemistry laureates recognised since 1901.
The Nobel Committee said the researchers had won the award for "the development of super-resolved fluorescence microscopy".
Profs Betzig and Moerner are US citizens, while Prof Hell is German.


Committee chair Prof Sven Lidin, a materials chemist from Lunds University, said "the work of the laureates has made it possible to study molecular processes in real time".

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Re: Microscope work wins Nobel Prize in Chemistry

#2  Postby Calilasseia » Oct 09, 2014 11:33 am

For those who need more on this, and why it's such a piece of absolute genius, here's an account I gave elsewhere, but which is apposite to reproduce here.

Heading off to the Official Nobel Prize website, and downloading the PDF covering the scientific background (document can be found here), we learn that the problem chosen as the focus of research by the latest Nobel Laureates, centres upon the well known limits that apply to any attempt to use electromagnetic waves for imaging. Simply put, it is impossible to resolve details, under normal circumstances, using an electromagnetic illumination source, when those details are closer together than L/2, where L is the wavelength of the electromagnetic radiation being used. This, of course, was the original motivation for moving to electron microscopy, because electron beams are associated with a far smaller value of L. In detail, visible light has a mean wavelength of around 500 nanometres, so details smaller than this cannot be resolved with a conventional light microscope, whereas the wavelength associated with an electron beam is of the order of 500 picometres - fully 1,000 times smaller - which means that an electron microscope has a resolving power that comes pretty close to that needed to resolve individual molecules of certain organic chemicals.

However, the electron microscope has a problem associated with it. Namely, that its superb resolving power is only usable upon specimens housed in a vacuum. As a corollary, it's impossible to examine living cells with an electron microscope, because those cells would pretty quickly stop living, when subjected to the conditions inside an electron microscope (ambient pressure 10-9 bar), and that's before we take into account that the electrons being used to illuminate the target are being accelerated by a potential difference of around a million volts. Though I'm minded to note at this juncture, that Tardigrades provided a surprise by surviving those conditions, and indeed, have since been determined to be capable of surviving in outer space for up to 30 days at a time, but I digress.

So, given that visible light is currently the only means of illuminating living cells, the researchers asked themselves, how they could solve the problem of being unable to resolve details smaller than L/2, a limit known as the Abbe Diffraction limit.

The authors then discuss, at some length, a number of past methods aimed at overcoming this limit, and the physics and mathematics involved, before moving on to their own technique. Which involves precise excitation of fluorescent molecules, in such a manner that their spatial position can be determined in a manner overcoming the Abbe Diffraction Limit. Mathematically, if α is half the aperture angle under which a point source of light is observed, and n is the refractive index of the medium through which the light is travelling, then the Abbe Difffraction Limit is given by :

δxmin = L/2n sin(α)

However, the experimenters discovered a neat trick. If they illuminate their fluorescent source with one, low-intensity laser, and simultaneously illuminate the same source with a second beam, of much higher intensity, and red-shifted with respect to the first beam, the resulting expression for δxmin in this setup becomes instead, the following expression:

δxmin = L/2Rn sin(α)

where R is an additional factor of the form:


where Isat is the saturation depletion intensity of the first beam, and I0 is the intensity of the second beam. As I0 increases, δxmin shrinks below the Abbe Diffraction Limit arbitrarily. The location of the flourescent spot and its spread therefore approaches, in the limit, the value of the Dirac Delta Function, which means that the spot's position is located to virtually arbitrary precision.

The beauty of this system, of course, is that the target can be any living cell, either in aqueous solution or growing on a suitable culture medium. By binding the molecule known as Green Fluorescent Protein (GFP for short) to target molecules in the cell, and illuminating them using this method, fine resolution detail can be determined in the living cell at a resolution that could match the resolution of an electron microscope.

But, even better still, by binding GFP to the right target molecules, the reaction chemistry of those target molecules can be rendered observable in real time, and at aribtrarily high magnification. Modern lasers can produce photon pulses separated by time periods as small as 10-15 seconds or better, and by using these pulses to track the changes in position of the GFP probes, the movement and the reaction chemistry of the target molecules to which the GFP has been attached, can be followed in real time, the data being logged by a fast computer as it is received via a CCD chip. The usual Fast Fourier Transform analysis can be used to filter out any residual noise, and deconvolution techniques used to resolve the data even further.

Now since it takes time for the Nobel Committee to announce their deliberations, and in this case, the work being awarded the prize dates back to 2004, we have now had, for ten years, a microscopy technique allowing us to analyse, in real time, the chemistry of the living cell at the molecular level.

This paper, not to put too fine a point on it, is a work of exquisite genius. These scientists found a way of circumventing what was originally considered to be an inviolable physical limit, by clever use of the spatial information provided by excitation generated photons, and the generation thereof in a manner precisely controlling the spatial origin thereof. In effect, the researchers have overcome the Abbe Diffraction Limit, by abandoning reliance upon arbitrarily generated photons from a simple radiant light source, and instead, generating photons to order in precise positions within a 2×2 array of "pixels" within the specimen, within a precisely defined focal plane. Because they are able to determine in advance the positions where they are generating the excited photons, and are able to do this within the sample 1015 times per second, the researchers are effectively able to "raster scan" their 2×2 "pixel grid" at extremely high speed, and track motion of molecules, including motion arising from chemical reactions.

This is how you win Nobel Prizes. Not by throwing your hands in the air, and declaring that magic and a magic man must be needed because you can't understand what's actually happening. Another stupendous victory for testable natural processes.
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Re: Microscope work wins Nobel Prize in Chemistry

#3  Postby Animavore » Oct 24, 2014 6:34 pm

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