Nature 466 , 647–651 (29 July 2010)

Miysaka Lab.Keisuke Yamada

Alexandros Pertsinidis, Yunxiang Zhang & Steven Chu

Contents

I. Introduction

II. Experiment

Sample & Setup

III. Results & Discussion

The separation between differently coloured fluorescent molecules

IV. Coclusion

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

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)≦

STED

+=

DMDM

Excitation laserSTED laser

The diameter fluorescence spot is decreased and get better resolution power

detector

Excitation spot STED spot

SNOM

~100 nm

SNOM is a kind of Scanning Probe Microscope using optical fiber as probe.

PALM/STORM

生物物理 50 （ 4 ）， 174-179 藤田克昌

Super-resolution with wide-field microscope

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

Sample

glass glasssilan

glasssilanPEG

glass

PEGsilan

biotin

coated glass

BSA Streptavidin biotin-PEG beads DNA

silanize coated with PEG coated with biotin

100 m

Setup

Closed-Loop feedback system

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

Sample

Laser Optical Tweezers

Polystyrene beadmeasurement

Locked by feedback system

Measuring the position of the Alexa 647and using the mapping calibration

Result

0.77 nm spatial resolution is achieved

a

b

c

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