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Nature 466 , 647–651 (29 July 2010)
Miysaka Lab.Keisuke Yamada
Alexandros Pertsinidis, Yunxiang Zhang & Steven Chu
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Contents
I. Introduction
II. Experiment
Sample & Setup
III. Results & Discussion
The separation between differently coloured fluorescent molecules
IV. Coclusion
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Optical detection
Optical microscope
A type of microscope which uses visible light and a system of lenses to magnify images of small samples
・ Disadvantage diffraction-limited optics ( 回折限界 ) QuickTime˛ Ç∆
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limiting the resolution of the microscopy(~200 nm)
・ Advantage nondestructive, noncontact observe inside of a sample
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δ =0.61× λNA
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δNA : numerical aperture
λ : Wavelength
: resolution
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Super-Resolving Microscopy
・ STimulated Emission Depletion (STED) Microscopy
・ Scanning Near Field Optical Microscopy (SNOM)
・ Photoactivated Localization Microscopy (PALM)
・ STochastic Optical Reconstruction Microscopy(STORM)
・ To identify the structure of a biological molecule・ Nanoscale property in materials can be evaluated.
Example of the method
The resolution power is higer than the optical microscope ( 100 nm)≦
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STED
+=
DMDM
Excitation laserSTED laser
The diameter fluorescence spot is decreased and get better resolution power
detector
Excitation spot STED spot
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SNOM
~100 nm
SNOM is a kind of Scanning Probe Microscope using optical fiber as probe.
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PALM/STORM
生物物理 50 ( 4 ), 174-179 藤田克昌
Super-resolution with wide-field microscope
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Motivation
To know about the structure of multisubunit biological complexesin biologically relevant environment
To improve the spatial resolution utilizing closed-loop feedback system
The spatial resolution is limited by noise and systematic effects that include drift and vibration of the measurement apparatus
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Sample
glass glasssilan
glasssilanPEG
glass
PEGsilan
biotin
coated glass
BSA Streptavidin biotin-PEG beads DNA
silanize coated with PEG coated with biotin
100 m
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Setup
Closed-Loop feedback system
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Result
The feedback systems controll the position・ By locking the pinhole position in each image, they avoid the effect of the apparatus’ drift
・ By locking a Cy3 molecule and kept same the position, they demonstrate 0.7 nm molecule to molecule reproducibitliy
320 nm
3.2 nm
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Sample
Laser Optical Tweezers
Polystyrene beadmeasurement
Locked by feedback system
Measuring the position of the Alexa 647and using the mapping calibration
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Result
0.77 nm spatial resolution is achieved
a
b
c
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Conclusion
・ The spatial resolution of 0.77 nm is attained for the optical microscope.
・ The Closed-roop feedback system could avoid vibration of the measurement apparatus and the combined effects of the relay imaging optics and CCD array.
・ The closed-loop control and registration could become essential concepts in design of future sub-10-nm optical lithography tools