Fourier-transform coherent anti-Stokes Raman scattering microscopy Jennifer P. Ogilvie et al. Opt....

Fourier-transform coherent anti-Stokes Raman scattering microscopy

Jennifer P. Ogilvie et al. Opt. Lett. 31, 480 (2006)

Kazuya MORI

MIYASAKA Lab.

Contents

Introduction ・ Raman Scattering ・ Coherent Anti-Stokes Raman Scattering (CARS) ・ Nonresonant background problem ・ Motivation Fourier-transform CARS (FTCARS) method Experimental Setup using a femtosecond laser Results and Discussion ・ CARS imaging of polystyrene bead Summary

http://www.nanophoton.jp/raman/raman-11.html

microscopy

Molecular vibrationAnti-Stokes Raman Scatteringωvib

Stokes Raman Scattering

Rayleigh scattering

Incident light

ν0

ν0+ωvib

ν0

ν0-ωvib

ωvib

Virtual state

+Raman Scattering

Stokes Anti-Stokes

Energy diagram

Raman Scattering

Introduction

excellent ”molecular fingerprint” for their identification

分子振動

ラマン散乱

nucleic acid protein lipid Raman spectra

Introduction

Coherent Anti-Stokes Raman Scattering (CARS)

But Raman scattering is・・・・ very weak signal (~10-6 of incident

radiation)・ difficult to separate from fluorescence

Molecular vibration

ωvib

ω1

beam2

beam1

ωvibω2

ω3= 2ω1 – ω2

(pump, probe)

(stokes)

ω1

ω1

ω2

ω3

・ significant signal enhancement over Raman scattering・ Anti-Stokes field → fluorescence-free

Nonresonant (NR) background Problem

Introduction

ωvib ωvib

ω’vib

Raman　 spectra of polystyrene beads conventional　 Raman microscope CARS microscope

NR background signal diagram

Near two-photon resonance

far from two-photon resonance

low resolution （ 30cm-

1）and spectral shift

非共鳴

M. Hashimoto et. al. Opt. Lett. 25, 1768-1770 (2000)

Motivation

NR background Limited fingerprint region imaging Complex setup

Problem

a new method for CARS microscopy with high resolution using a single femtosecond pulse laser

Single femtosecond pulse laser

Fourier-transform CARS (FTCARS) method

two femtosecond pulse

ωvib

time-domain

Time delay τ/ fs

vibrational polarization + ② Time-domain CARS signal振動分極

stokespump

probe

① ②

CARS

τ

delay

Fourier-transform CARS (FTCARS) method

Fourier-transform

frequency-domaintime-domain

Time delay / fs frequency / cm-1

CARS spectrafrequency-domain spectra =

NR background

Fourier-transform

vibrational polarization

＝Molecular vibration

)(tx

frequencies

dtetxX ti )()(

)(X

(electronic response)

dteXtx ti)()(

Experimental Setup

~820nm Pulse duration ~20fs

PC: prism compressor

LPF: long-pass filter(>780nm)

SPF: short-pass filter(<760nm)

BS: beam splitter

DBS: dichroic beam splitter

PMT: photomultiplier tube

fluorescence

CARS signal

delay

光電子増倍管

ω3= 2ω1 – ω2

ω1

ω2 Phase matching 2k1k2

k3

Conventional CARS CARS microscopy

ω3ω1

ω2(CARS)

Filter spatially-resolved

spectral filtering

Results and Discussion

HC

H2C

NR background

Sample

20μm polystyrene bead

in 2-propanol

time-domain

vibrational polarization

Time delay (fs)

ampl

itude

(a.

u.)

H3CHC CH3

CH3

2-propanolpolystyrene

Results and Discussion

FTCARS imaging

FTCARS offers high spectral resolution ~13cm-1

Scan

1005cm-1 → C-C stretching mode in polystyrene816cm-1 → C-C stretching mode in 2-propanol

Summary

A Fourier-transform technique for CARS microscopy that employs a single laser source and time-delay setup was proposed.

FTCARS method was demonstrated by spectrally imaging a polystyrene bead in 2-propanol.

FTCARS microscopy offers a compact optical setup and high spectral resolusion(～ 13cm-1).

ω2

Frequency / cm-1

femtosecond pulse

ω1

time/ fs

　 uncertainty principle 　 ΔEΔt~ħ

stokespump

probe

① ②

CARSω1ω2

Spontaneous Raman spectrum of polystyrene

http://riodb01.ibase.aist.go.jp/sdbs/cgi-bin/direct_frame_top.cgi

HC

H2C

Sample

Sample

Spontaneous Raman spectrum of 2-propanol

http://riodb01.ibase.aist.go.jp/sdbs/cgi-bin/direct_frame_top.cgi

H3CHC CH3

CH3

http://www.bangslabs.com/cgi-bin/PSGFind.pl?return=6.2&code=FC07F

Fluorescent dye

Sample

photomultiplier tube　（光電子増倍管）

http://ja.wikipedia.org/wiki/%E7%94%BB%E5%83%8F:Photomultipliertube.png