Download Surface Plasmons

Nanoplasmonic
Biomolecular Imaging
(奈米電漿子生物分子影像)
Shean-Jen Chen (陳顯禎)
3-30-2006
Adaptive Photonics Lab, NCKU
- Nanoplasmonic Biosensing & Molecular Imaging
- Adaptive Optics for Vision Science
Outlines
- Surface Plasmons & Particle Plasmons
- Label-free Nano-imaging : Surface Plasmon Resonance
(SPR) Microscopy
- Amplified Optical Near-field : Metal-tip Plasmon-enhanced
Near Field Scanning Optical Microscope (NSOM) for
Fluorescent or Raman Molecular Image
- Breaking Diffraction Limit : Nanoplasmonic Structured
Metalayer
- Conclusions
Adaptive Photonics Lab, NCKU
Optical Microscopy
Advantages:
- Intuitive image interpretation
- Applicable to samples in natural
environment
- In general non-destructive
- Easy to use
Disadvantages:
- Abbe diffraction limit
- Large sample area exposed
to illumination light
 min
0.61 

NA


2
if NA  1.2
Total Internal Reflection Fluorescence
Microscopy (TIRFM)
- Intensity Measurement:
Controlling the depth resolution
via changing incident angle or
wavelength

2
2
dp  1/  
n1 sin 2 1  n2
2


1 2
Evanescent Wave
Filter out scattering light
“ What bother the TIRF ?” It can provide a better depth resolution
than confocal microscopy, especially when it comes to selecting
fluorescent molecules close to the biosensing surface.
Adaptive Photonics Lab, NCKU
Outlines
- Surface Plasmons & Particle Plasmons
- Label-free Nano-imaging : Surface Plasmon Resonance
(SPR) Microscopy
- Amplified Optical Near-field : Metal-tip Plasmon-enhanced
Near Field Scanning Optical Microscope (NSOM) for
Fluorescent or Raman Molecular Image
- Breaking the Diffraction Limit : Nanoplasmonic Structured
Metamaterial
- Conclusions
Adaptive Photonics Lab, NCKU
Surface Plasmon Resonance (SPR) Biosensors
(Kretschmann-Raether Configuration)
Slide
Au film
SAM
Receptors
Ligands
Prism
P-wave light
Detector
1
θ
0.9
Slide
Au film
O-ring
0.8
0.7
Flow cell
TE-cooler
SAM out
Ligand
Receptor
out
out
Intensity
Receptor
Ligand
SAM ininin
0.6
0.5
0.4
0.3
0.2
SPR condition:
k x  k0 ε0 sinθ  k sp
k sp  k0
ε 1ε 2
ε 1 ε 2
0.1
0
53
54
55
56
Coupling Angle
57
58
59
(Surface plasmon wave at semi-infinite structure)
Adaptive Photonics Lab, NCKU
Advantages of SPR Biosensing
Theoretical sensitivity
• Label-free sample
• Real-time biomolecular
interaction analysis (BIA)
• Kinetic study
• Characterize and quantify
biomolecular interaction
• High sensitivity
(~ 1 pg/mm2)
• Potential of high
throughput screening
(Resolution unit: RIU; λ= 632.8 nm)
Spectrum
Intensity
Detection
approach
Angular
Wavelength
Intensity
Phase
Prism
coupler
5 x 10-7
2 x 10-5
5 x 10-5
2 x 10-7
Grating
coupler
2 x 10-6
6 x 10-5
2 x 10-4
8 x 10-7
1. ATR coupler: BK7 / Au (48 nm) / analyze (1.32).
2. Grating coupler: Pitch of 800 nm & depth of 70 nm.
3. Angular resolution is 1 x 10-4 deg.
4. Wavelength is 0.02 nm.
5. Intensity resolution is 0.2 %.
6. Phase resolution is π/200.
Adaptive Photonics Lab, NCKU
Intensity and Phase Variation at SPR
- Reflection relationship:
p
R  r012
2
r01p  r12p exp  2ik z1d 

1  r01p r12p exp  2ik z1d 
2
p
  tan 1  r012
  Im  r012p  Re  r012p 
SPR curve /Phase -Prism/Au/Protein/Water
SPR curve /Magnitude -Prism/Au/Protein/Water
4
1
s-wave
0.9
3
0.8
p-wave
Phase shift (radian)
Reflectivity
0.7
0.6
0.5
0.4
0.3
2
18nm
1
0
-1
-2
0.2
s-wave
-3
0.1
0
55
6nm
p-wave
6nm
60
65
70
18nm
75
80
85
-4
55
60
65
70
75
80
85
Incident Angle (degree)
Incident Angle (degree)
- Phase variation analysis:
r  r exp  i 
Im
r'
r'
 '  Im  r ' r 
tan 1  r ' r    2  n
b
Re
Adaptive Photonics Lab, NCKU
Particle Plasmon Resonance
(D.A. Schultz, 2003)
Adaptive Photonics Lab, NCKU
Plasmonic Biosensors
4122.36
4122.
3000
30
- Particle plasmons
- Gap mode
Z
0
2000
0.
0.1
0.2
0.3
X
0.4
0.5
4122.36
0.21
0.6
Y cut = 0
0.15
0.1
3000
2000
Y
Z
θ
0
1000
-0.1
0.21
0
0.1
0.2
0.3
X
0.4
0.5
-0.15
0.6
l
D
d
→ To increase local EM field (about 105 times)
→ To increase sensitivity (less than 1 pg/mm2)
G.-Y. Lin et al., Proc. SPIE 6095 (2006).
20
10
1000
-0.1
-0.15
0
Z
0.1
Y
Four Plasmonic Effects:
- Surface plasmons
- Inter-particle coupling
0.15
50 nm
Adaptive Photonics Lab, NCKU
Nanoparticle-enhanced Plasmonic Biosensors
• Excitation of surface plasmons and particle plasmons
Au film
• Locally enhanced EM fields
Glass
• Providing more sensitive biosensors
Photodiode 1
TE-wave
He-Ne Laser
Polarizer
Half-wave plate
Receptor
Ligand
Flow in
TM-wave Photodiode 2
Wollaston prism
θ
Prism
Au thin film ~40 nm
Au:SiO2 composite film ~15 nm
Silane
O-ring
Flow cell
Flow out
• Grain size 4.0 nm &
interval 2.0 nm
• Not analyte-tagged
nanoparticles
• Not synthesizing
W. P. Hu et al., Biosensors & Bioelectronics 19 (2004) 1465.
S.-J. Chen et al., U.S. Patent Pending No. 10/660833, 2003.
Adaptive Photonics Lab, NCKU
Advanced Plasmonic Biosensing
What is Next?
Biomolecular Imaging with Plasmonic Effects
Adaptive Photonics Lab, NCKU
Outlines
- Surface Plasmons & Particle Plasmons
- Label-free Nano-imaging : Surface Plasmon Resonance
(SPR) Microscopy
- Amplified Optical Near-field : Metal-tip Plasmon-enhanced
Near Field Scanning Optical Microscope (NSOM) for
Fluorescent or Raman Molecular Image
- Breaking the Diffraction Limit : Nanoplasmonic Structured
Metamaterial
- Conclusions
Adaptive Photonics Lab, NCKU
Common-path Phase-Shift Interferometry
SPR Imaging
- Long-term stability to reject external disturbances
- Easy aligned and compact system
CCD
Analyzer
Beam
Splitter
Phase-shifting
Image
Liquid Lens
Laser
Polarizer
Crystal
Beam
Expender
θ
Prism
Phase
variation
Analyzer
Photodiode
Cr (3nm)
Au (46nm)
Sample
Mapping
to CCD
Flow Cell
TE Cooler
S.-J. Chen et al., Journal of Biomedical Optics 10 (2005) 034005.
Adaptive Photonics Lab, NCKU
15mer DNA SPR Phase Image
- Five-step Phase-shift Interferometry
 500m
 2I 2  I 4  

2
I

I

I
 3 5 1
 ( x, y )  tan 1 
0
1/2π
π
3/2π
2π
- Phase Reconstruction
0.8
DNA
phase ()
0.6
DNA
0.4
0.2
0
-0.2
Y.-T. Su et al., Optics Letters 30 (2005) 1488.
0
100
200
pixel
300
400
Adaptive Photonics Lab, NCKU
SPR Microscopy
- SPR Phase Microscope for Living Cell Membrane Images
with No Fluorescent Labels
- Plasmon-enhanced TIR Fluorescence Microscope for
Dynamic Living Cell Membrane Images
Detecctor
Filter
Incident light to
excite SPs & PPs
Collecting lens
Fluorescence
‘Surface Plasmons’
&
‘Particle Plasmons’
Prism
Metal thin film
Metal nanopatricles
Antibody
Antigen
Fluorescencemt label
Adaptive Photonics Lab, NCKU
Cell Membrane Imaging
Melanoma-GFP-tagged TM cell
TIRFM
Plasmon-enhanced TIRFM
400
- The enhancement of fluorescence
is observed apparently btw the
two images. The experimental
results show that the
fluorescence intensity can be
enhanced about 3.0 fold.
SPR
TIR
350
300
Intensity(a.u.)
250
200
150
100
50
0
0
20
40
60
80
100
pixel
120
140
L.-Y. He et al., Proc. SPIE 6088 (2006).
160
180
- GFP-tagged TM on the melanoma
cell membrane near the chip
surface is excited by the
evanescent wave for TIR or
surface plasmon wave for SPR;
200
- Because of the variant distance
btw the cell membrane protein TM
and the collagen-coated surface,
different surface plasmon effects
can be applied to interpret the
phenomenon of fluorescence
emission or quenching.
Adaptive Photonics Lab, NCKU
Lateral Resolution Limited by
Propagation Length of Surface Plasmon Wave
Ag
Al
Propagation Length @ λ=630nm
Ag = 19μm; Al = 1 μm
A Goldfish Glial Cell
Interference Reflection Microscope
Ag
Al
K.-F. Giebel et al., Biophysical Journal 76 (1999) 509.
SPR Intensity Microscope
Adaptive Photonics Lab, NCKU
Outlines
- Surface Plasmons & Particle Plasmons
- Label-free Nano-imaging : Surface Plasmon Resonance
(SPR) Microscopy
- Amplified Optical Near-field : Metal-tip Plasmon-enhanced
Near Field Scanning Optical Microscope (NSOM) for
Fluorescent or Raman Molecular Image
- Breaking the Diffraction Limit : Nanoplasmonic Structured
Metamaterial
- Conclusions
Adaptive Photonics Lab, NCKU
Near-field Scanning Optical Microscope
(NSOM)
Operation:
• Confined by a metal aperture
• Within short distance beyond
the screen.
Evanescent Wave and Nanoscanning Tip Techniques to
Break “Diffraction Limit”
• Lateral spatial resolution: ~20nm
• Longitudinal spatial resolution: ~50nm
Applications:
•
•
•
•
Single molecule to cell detection
Nanolithography
Super-resolution data storage
Near-field optical interaction on
nanoparticles, nanoclusters,
and localized surface plasmon.
• Nanophotonics
• Surface photochemistry
Adaptive Photonics Lab, NCKU
Single-molecule Detection
Liquid operation of NSOM opens the way to
directly visualise and quantify the size and
composition of membrane domains, like lipid
rafts, in solution. Fluorescence image of a
dendritic cell in buffer solution collected in
confocal mode (A) and NSOM mode (B).
FEBS Letters 573 (2004) 6.
Journal of Cell Science 114 (2001) 4153.
Single molecule detection on cells by NSOM. This
figure shows a 40 nm optical resolution near-field
‘zoom-in’ on the indicated area (3.2 mm2) in the
bright-field image of a fibroblast expressing LFA-1GFP. GFP excitation is accomplished using 488 nm
light (Ar-Kr laser line) linearly polarized along 90°.
Apertureless NSOM
In apertureless near-field scanning optical microscopy (ANSOM), the probe
vibration is often used to increase the detected signal that can be detected at
by a lock-in amplifier. The realistic model of ANSOM should take into account
the scan of the probe as well as the probe vibration and the material properties.
R. Fikri et al., Optics Letters 28 (2003) 2147.
Adaptive Photonics Lab, NCKU
Surface-Enhanced Raman Scattering (SERS)
• Amplified local EM fields
• To study protein conformational change
• To investigate biomolecular structures
Incident light
to excite SPR
θ >θ critical
Reflected beam
θ
Prism
Au thin film ~48 nm
Au nanopatricles
SiO2 (400~600 nm)
O-ring
Collecting lens
Flow cell
Receptor
Ligand
Flow in
Flow out
C. L. Lei et al., Mater. Sci. Eng. B 32 (1995) 39.
To optical spectrometer
Adaptive Photonics Lab, NCKU
Metal tip-enhanced NSOM for Fluorescent
Molecular Image
- Enhance local EM field & improve
detecting signal
- High resolution image for fluorescent
DNA samples
H. Frey, PRL 93 (2004) 200801.
DNA Fluorescent Image
Adaptive Photonics Lab, NCKU
Metal tip-enhanced Raman Spectroscopy
N. Hayazawa et al., JAP 92 (2002) 6983.
Outlines
- Surface Plasmons & Particle Plasmons
- Label-free Nano-imaging : Surface Plasmon Resonance
(SPR) Microscopy
- Amplified Optical Near-field : Metal-tip Plasmon-enhanced
Near Field Scanning Optical Microscope (NSOM) for
Fluorescent or Raman Molecular Image
- Breaking Diffraction Limit : Nanoplasmonic Structured
Metamaterial
- Conclusions
Adaptive Photonics Lab, NCKU
Stimulated Emission Depletion (STED)
Microscopy
The role of the STED beam is to induce
the transition L2 L3 by stimulated
emission and to deplete the excited
fluorescence.
Excitation pulses are followed by stimulated
emission depletion pulses for fluorescence
inhibition. After passing dichroic mirrors and
emission filters, fluorescence is detected through a
confocal pinhole by a counting photodiode.
PNAS 97 (2000) 8207.
Adaptive Photonics Lab, NCKU
Comparison btw Confocal & STED
Reducing the fluorescence focal spot size
to far below the diffraction limit: (a) spot
of a confocal microscope (left) compared
with that in a STED microscope (right)
utilizing a y-oriented intensity valley for
STED (upper right insert, not to scale)
squeezing the spot in the x direction to 16
nm width.
(b) The average focal spot
size (squares) decreases with the STED
intensity following a square-root law. Insert
(right) discloses the histogram of the
measured spot sizes rendering the 26 nm
average FWHM.
PRL 94 (2005) 143903.
Adaptive Photonics Lab, NCKU
Negative Refractive Index
In 1968, Veselago first considered the case of a medium that had both
negative dielectric permittivity and negative magnetic permeability.
Nature 420 (2002) 119.
Adaptive Photonics Lab, NCKU
Negative Refraction Makes a Perfect Lens
Flat Slab
Object
Real Image
A negative refractive index medium
bends light to a negative angle with the
surface normal. Light formerly diverging
from a point source is set in reverse and
converges back to a point.
J. B. Pendry, PRL 85 (2000) 3699.
Adaptive Photonics Lab, NCKU
Microwave Imaging by Flat Lans using
Negative Refraction
Metal Nanostructure with square
copper split ring resonators and
copper wire strips
R. A. Shelby et al., Science 292 (2001) 77.
Photonic Crystal at Band-gap Edge
for 9.3 GHz
The images can be observed only in a narrow
frequency range, between 9.0 and 9.4 GHz.
P. V. Parimi et al., Nature 426 (2003) 404.
Adaptive Photonics Lab, NCKU
Sub-Diffraction-Limited Optical Imaging with
a Silver Superlens
N. Fang et al., Science 534 (2005) 308.
Enhanced Lateral Resolution by Control of the Size
& Distribution of Metal Nanoparticles
Enhanced the Readout
Signal of Super-RENS Disc
J.-N. Yih et al., Applied Optics 44 (2005) 3001.
- Break diffraction limit &
improve single particle
detection without near-field tip
- Lateral spatial resolution:
~100nm
Adaptive Photonics Lab, NCKU
Coupled Waveguide-surface Plasmon
Resonance Biosensors Constructed with
Sub-wavelength Grating
White beam & normal incident (TM)
Metal layer
(δ = 60nm)
grating layer
(δ = 25nm)
guided-layer
( d = 250 nm)
0.5Λ
0.9
biolayer: 10nm / 1.46
0.8
0.7
n2 / Ta2O5
n3 / BK7
substrate
(1000μm)
Red line: without a bio-layer
Blue line: with a bio-layer
1
n1 / H2O
Intensity
Λ=380nm
0.6
0.5
0.4
0.3
0.2
0.1
0
590
21.2 nm
600
610
620
630
640
650
660
Incident wavelength (nm)

The reflectivity spectrum of CWSPR biosensors with sub-wavelength grating
 Localized surface plasmon mode and waveguide mode
 Sharper dip-width
 Enhance sensitivity
Adaptive Photonics Lab, NCKU
EM Wave transport below the Diffraction Limit
in Metal Particle Plasmon Waveguides
(S. A. Maier et al., 2003)
Adaptive Photonics Lab, NCKU
Anticipated Research Outcomes
• Establish the immobilization of biomolecules
• Build the real-time portable SPR and SERS metrology systems
• Using plasmonic biosensors to achieve the direct detection of drug
• Conformational & Structural information from CWSPR & SERS
• A total solution of the operation of advanced plasmonic biosensing
 Single Bio-molecule Detection for BIA
 Molecular Scale Imaging of Living Cell
Adaptive Photonics Lab, NCKU