Stochastic Optical Reconstruction Microscopy (STORM)

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    02-Jan-2016
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Stochastic Optical Reconstruction Microscopy (STORM). Finding Out the Position of a Molecule. FIONA. σ  σ PSF / N 1/2. Thompson at al., Biophys. J. , 2002. Yildiz et al., Science , 2003. Localization by sequential photobleaching. or QD blinking. A few molecules. - PowerPoint PPT Presentation

Transcript of Stochastic Optical Reconstruction Microscopy (STORM)

  • Stochastic Optical Reconstruction Microscopy (STORM)

  • Finding Out the Position of a Molecule PSF / N1/2Yildiz et al., Science, 2003Gordon et al., PNAS, 2004Lagerholm et al., Biophys. J., 2006FIONALocalization by sequential photobleachingThompson at al., Biophys. J., 2002Qu et al., PNAS, 2004Lydke et al., Optics Exp., 2005or QD blinkingA few molecules

  • Many Molecules

  • The principle of STORM

  • The principle of STORM

  • The principle of STORM

  • The principle of STORM

  • The principle of STORM

  • The principle of STORM

  • The principle of STORM

  • The principle of STORM

  • The principle of STORM

  • The principle of STORM

  • The principle of STORM

  • The principle of STORM

  • A Photo-switchable ProbeActivationDeactivationImaging laser (657 nm)Activation laser (532 nm)ActivatorReporter6000 photons

  • More ColorsActivator absorption (nm)Reporter emission (nm)Alexa 405Cy2Cy3Cy5Cy5.5Cy7665690775550490400Alexa 647Activation pulsesActivation pulsesActivation pulsesFluorescenceFluorescenceFluorescence405 nm532457Bates et al, Science 317, 1749 1753 (2007)

  • 5 mB-SC-1 cell, Microtubules stained with anti- tubulinCy3 / Alexa 647 secondary antibody

  • 5 mBates et al, Science 317, 1749 1753 (2007)

  • 5 m

  • 5 m Cy3 / Alexa 647: Clathrin Cy2 / Alexa 647: MicrotubuleBates et al, Science 317, 1749 1753 (2007)

  • 1 m

  • 200 nmAvg = 172 nm

  • Class evaluationWhat was the most interesting thing you learned in class today?

    2. What are you confused about?

    3. Related to todays subject, what would you like to know more about?

    4. Any helpful comments.Answer, and turn in at the end of class.

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