imaging through scattering - micronanoconference · ivo vellekoop biomedical imaging with light 0.1...
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Imaging through scattering
Ivo Vellekoop
Ivo Vellekoop
Biomedical imaging with light
0.1 mm 1 mm 10 mm 100 mm
2
2
< 50 nm resolution
~ 1 µm resolution
~ 0.5mm resolution
Ivo Vellekoop
More than tissue?
Insert your favorite specimen:
• Integrated circuits
• Microfluidic systems
• Organ on a chip
• Opaque catalysts
• Powders
All ideas are welcome!
Ivo Vellekoop
Newtonian Ray Optics
4
lens
All microscopes rely on Newtonian ray optics basic principle has not changed in 400 years:
Ivo Vellekoop
Scattering
In tissue light does not propagate along straight rays
Ivo Vellekoop
Biomedical imaging with light
0.1 mm 1 mm 10 mm 100 mm
6
2
Paradigm 1: microscopy
Paradigm 2: diffuse tomography
full body microscopy of an intact mouse
complete full mouse brain at dendritic resolution Dream on …
… or get to work!
Ivo Vellekoop
Looking through scattering
All microscopic approaches are aimed at: • Reducing scattering:
– Chemical clearing – Skull thinning – Multi-photon microscopy
• Rejecting scattered light – Optical coherence tomography – Confocal microscopy – Multi-photon microscopy
How about using scattered light?
Ivo Vellekoop
From speckle to focus
Vellekoop & Mosk. Opt. Lett. 31, 2309 (2007)
spatial light modulator (LCD)
Ivo Vellekoop
Before
detector
total field in target
Ivo Vellekoop
After
detector
total field in target
Ivo Vellekoop
Ivo Vellekoop
Focusing better than with a lens
Vellekoop, Lagendijk, Mosk, Nature Photonics 2010.
Ivo Vellekoop
Focusing light inside
Vellekoop, van Putten, Lagendijk & Mosk. Opt. Expr. 16, 67 (2008)
Ivo Vellekoop
Focus + scanning + fluorescence
Focus + scanning + fluorescence = microscope
Vellekoop & Aegerter. Opt. Lett. 35, 1245 (2010)
Ivo Vellekoop
The optical memory effect
Maximum scan angle (HWHM): α ≈ 0.24 λ/ L
Ivo Vellekoop
Imaging results
Used scattered light for imaging!
Ivo Vellekoop
Resolution
300 nm resolution
with multiply scattered light!
HWHM:300 nm
Vellekoop & Aegerter. Opt. Lett. 35, 1245 (2010)
Ivo Vellekoop
Inside?
How do you get the initial focus?
Need some form of feedback
We are not allowed to peek
Ivo Vellekoop
Now we have many options:
• Iterative – 1-photon fluorescence, ultrasound tagging,
photo acoustic effect, non-linear PA, 2-photon fluorescence, …
• Phase conjugation – Ultrasound tagging (TRUE)
– Movement tagging (TRACK)
– Fluorescence, 2nd harmonic, …
Ivo Vellekoop
Imaging through a mouse skull
Park, Sun & Cui. PNAS 2015
Feedback comes from strength of 2-photon signal!
Park, Sun & Cui. PNAS 2015
Ivo Vellekoop
“High-resolution in vivo imaging of mouse brain through the intact skull” Park, Sun & Cui. PNAS 112, 9236 (2015) [scale bar = 5 µm]
Ivo Vellekoop
Ivo Vellekoop
Are we already there?
Challenge: - Problem for looking through very thin scattering
layer more or less solved
- How about looking inside a thick scattering material? - Focusing possible. - Need ways to scan!
- fundamental understanding of wave correlations inside forward scattering materials (tissue)
- experimental validation - numerical validation
Ivo Vellekoop
Anisotropic scattering
Ivo Vellekoop
Memory effects
Gerwin Osnabrugge
Tilt/Tilt memory effect (thin layers only)
L
Feng et al. PRL 1988.
Shift/Shift memory effect (tissue-like scattering only)
Judkewitz et al. Nat. Phys. 2015.
Ivo Vellekoop
Combining shift and tilt
Feng et al. PRL 1988. Judkewitz et al. Nat. Phys. 2015. Osnabrugge et. al. (in preperation)
Ivo Vellekoop
Joint memory effect
Experiment Simplified analytical model
Osnabrugge et. al. (in preperation)
Ivo Vellekoop
Combining shift and tilt
Osnabrugge et. al. (in preperation)
Rotate one way, shift the other way Maximizing overlap ‘volume’
Ivo Vellekoop
How does it look inside the medium?
Ivo Vellekoop
Shift memory effect inside?
Ivo Vellekoop
About the simulations
Pack
er e
t al
. Nat
ure
Met
ho
ds
6, 2
10
2 (
20
12
)
Ivo Vellekoop
Validation of simulation method
Osnabrugge, Leedumrongwatthanakun, Vellekoop. J. Comp. Physics 2016.
Pseudospectral time domain: More accurate? slower
Our method: Alwayst fast & extremely accurate
Ivo Vellekoop
Almost perfect focus
Ivo Vellekoop
Aberrated focus
Ivo Vellekoop
• No ballistic component distinguishable
• No homogeneous diffuse background
• All features are sharp (speckle)
Ivo Vellekoop
Ivo Vellekoop
Ivo Vellekoop
Ordinary scanning
x [µm]
y [µ
m]
39
Ivo Vellekoop
Exploiting the memory effects
optimally (simulation)
40
x [µm]
y [µ
m]
Ivo Vellekoop
Conclusion
• Scattered light can be used for microscopic imaging
• Imaging through a thin slab of material is possible
• Imaging inside tissue still challenging, but progress:
– Anisotropic memory effect can be used
– Theoretical understanding of joint shift/tilt effects
– Extremely efficient & accurate simulation tools
– Promising results in simulation
Ivo Vellekoop
Biomedical Photonic Imaging (University of Twente)
Gerwin Osnabrugge
Saroch Leedumrongwatthanakun
Changhuei Yang lab (Caltech)
Benjamin Judkewitz lab (Charité Berlin)
Roarke Horstmeyer, Ioannis Papadopoulos
Allard Mosk (University of Utrecht)
Complex Photonics Systems (University of Twente)
Willem Vos, Ad Lagendijk
Christoph Aegerter lab (University of Zurich)
Acknowledgements
PhD and PD positions available
Ivo Vellekoop
Imagine…
Elke Vockenhuber
SCATTERING AND ABERRATIONS IN MICROSCOPY
Ivo Vellekoop
Aberrations
• Lenses – Spherical aberrations – Chromatic aberrations – Astigmatism
• Cover-glass • Misalignment • Sample
Ivo Vellekoop
Adaptive optics
Ray optics, smooth aberrations
Correct for: • System aberrations • Turbulent atmosphere • Lens of human eye • Cover slip thickness • …
Ivo Vellekoop
Aberrations vs scattering
Ray optics, smooth aberrations
Multi-path interference, wavelength-scale disorder speckle
Ivo Vellekoop
Atmosphere Paint
Ray optics Wave optics: diffraction, multiple scattering, interference
1 ray in = 1 ray out 1 ray in = multi-path interference
1 optical path length Distribution of optical path lengths -> diffusion in both space and time
Start with low-quality image Start with speckle
Ivo Vellekoop
Are scattering and aberrations fundamentally different things?
No: they are different ends of a spectrum of distortions
(like ice and steam are basically the same thing)
Ivo Vellekoop
Is this Adaptive Optics?
Absolutely: there is optics and there is adaptation
However: • For decades, the term Adaptive Optics has
been used far to narrowly, dealing mostly with aberrations in optical imaging systems
• Until 2007, a layer of paint was not considered to be an optical imaging system
Magic You can do the same without first
creating a focus!
Ivo Vellekoop
Raster scanning with unknown PSF
Bertolotti, van Putten, Blum, Lagendijk, Vos & Mosk, Nature 491, 232–234 (2012).
Ivo Vellekoop
How does the magic work?
• Raster scanning microscope
• With unknown PSF and unknown object
• Use statistical properties of PSF together with phase retrieval algorithm to recover image
Ivo Vellekoop
Single shot magic
Katz, Heidmann, Fink & Gigan, Nature Photonics 8, 784–790 (2014)
Ivo Vellekoop
Applications?
• Very cool ideas • For small objects far behind a thin screen
Applications for microscopy limited: – Through thin aberrating layer (skull) – Imaging inside eggs?
• Without memory effect, you need a different wavefront for every given scan point.
• This wavefront is a-priori unknown
Ivo Vellekoop
Novel memory effect
Judkewitz, Horstmeyer, Papadopoulos, Vellekoop & Yang, Nature Physics 2015.
Ivo Vellekoop
Exploiting the memory effect
THE MICROSCOPE OF THE FUTURE?
Ivo Vellekoop
Scanner
Future
Laser
Primary information: fluorescence signal
Secondary information: • wavefront distortion • OCT • background signal • sample movement/distortion
Smart Scanner
a-priori information: • system aberrations • memory-effect ranges • guide-star shape & distribution • refractive index models • recognizable features
Spatial Light Modulator
Ivo Vellekoop
Where are we now?
(based on Ntziachristos, V. Nature Methods, 7, 603–614 (2010))
1 MFP ~ 30 µm 1 TMFP ~ 1 mm
Paradigm 1: microscopy
Paradigm 2: diffuse tomography
Scattered light fluorescence microscopy
TROVE
TRUE
Brave new world: scattered light microscopy
full body microscopy of an intact mouse
Ivo Vellekoop
Conclusion
High-resolution imaging with scattered waves
A largely unexplored territory that requires a completely different way of thinking about imaging.
• Focus light anywhere if ‘only’ the correct wavefront is known
• First applications are already a fact.
Ivo Vellekoop
The Future
Microscopic endoscopy through a thin injection needle Full body microscopy of living animals Seeing through ‘walls’, fog, skin. Looking inside eggs, paint, composites, granular materials, IC’s, etc., etc.
Ivo Vellekoop
More reading
Review of feedback-based wavefront shaping:
• Vellekoop, Optics Express 23, 12189 (2015)
Review of digital optical phase conjugation:
• Horstmeyer et al., Nature Photonics (2015)
New memory effect
• Judkewitz et al., Nature Physics 11, 684 (2015)
Ivo Vellekoop
Time reversal of ultrasound encoded
light (TRUE)
• X. Xu et al. Nat. Photonics (2011).
• Y. M. Wang et al. Nat. Communications (2012). K. Si et al. Nat. Photonics (2012).
ultrasound lock-in 3D fluorescence microscopy
Ivo Vellekoop
TRUE Deep tissue fluorescence
microscopy
Deep-tissue focal fluorescence imaging with digitally time-reversed ultrasound-encoded light Ying Min Wang, Benjamin Judkewitz, Charles A. DiMarzio & Changhuei Yang Nature Communications 3, 928 (2012). Scale bars 50 μm.
Ivo Vellekoop
The problem
• US-assisted focusing (TRUE) is a great solution. Problem:
– Low resolution (~ 20 µm).
– Worse: contrast (~ 1:20)
• Contrast inversely proportional to number of modes in the target focus [Vellekoop & Mosk. Opt. Commun 281 (2008)]
Ivo Vellekoop
Solutions: • More pixels • Smaller ultrasound tagging volume • Longer wavelength • TROVE (Benjamin Judkewitz) / more efficient methods?
Not as bad as thought before: - intensity correlations result in more efficient focusing [Cui2016]
Ivo Vellekoop
Ivo Vellekoop
Example: wavefront-shaping in fatty tissue FDTD
71
Our novel approach FDTD
Computationally constructing transmission matrix of scattering tissue is becoming feasible
Ivo Vellekoop
Graphical representation
Re E
Im E
contribution of
segment 1 contribution of
segment 2
contribution of
segment N
Maximize Eb
Ivo Vellekoop
Algorithm
Adjust phase of individual segments
until contribution is in phase with total field
Ivo Vellekoop
Global maximum
before after
all the same phase
global maximum
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