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Characterising and improving image quality in Optical Coherence Tomography
and Elastography by means of optical beam shaping and simulations
Andrea Curatolo
BSc, MSc
This thesis is presented for the degree of Doctor of Philosophy
of The University of Western Australia School of Electrical, Electronic & Computer Engineering
January, 2017
THE UNIVERSITY OF WESTERN AUSTRALIA
ii
Abstract
Most diseases manifest themselves when symptoms affect the patient at the organ level, but
they originate at the molecular and cellular level, with clear signatures, such as changes to
morphology or stiffness. Understanding disease at such a level may permit earlier, more
specific diagnosis, and may improve targeted treatment strategies; however, it requires
imaging techniques that are sensitive and specific to such small variations in cell and tissue
micro-environment properties. Unfortunately, medical imaging technologies, capable of
performing such high-resolution imaging at depths of up to several millimetres in tissue,
are limited and need improvement.
One path to understanding the genesis and progression of disease, and to early disease
detection, lies in the advancement of such a microscopy technique. Optical coherence
tomography (OCT) holds much promise towards this end, as an optical, three-dimensional,
non-invasive, high-resolution imaging technique, with the ability to penetrate tissue,
reaching 2-3 mm below the surface. Optical coherence elastography (OCE), an extension
of OCT to image a sample’s three-dimensional stiffness distribution, can aid in this
scientific and clinical effort, by providing complementary information on the tissue
mechanical properties.
Nevertheless, the penetration depth and image contrast of OCT are fundamentally
limited by light scattering in biological tissue. In addition, OCE currently lacks the
resolution to visualise mechanical interactions at the cellular scale.
Characterisation and improvement of image quality in OCT and OCE are fundamental
to the translation of these techniques into reliable, non-invasive providers of biological
tissues’ microscopic structural and functional information, and for their range of
applications.
The first part of this thesis describes the methods used: firstly, to alter and improve
OCT and OCE image quality; secondly, to compare it in realistic and controlled turbid
tissue scenarios; and, thirdly, to analyse it. We use energy-efficient Bessel beams to alter
image quality, as a viable alternative to conventional focussing schemes using Gaussian
beams. We design novel structured phantoms to compare image quality in turbid tissue,
providing fine control of the optical, mechanical and structural properties, and aiding in the
benchmarking effort. We implement and use novel simulations of beam propagation and
image formation in turbid samples to analyse image quality and guide system development.
In the second part of this thesis, we employ the previously described tools to quantify
the effect that using energy-efficient Bessel beams has on OCT image quality in turbid
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tissue. We demonstrate that the Bessel beam’s increased depth of field comes at the
expense of both reduced peak OCT sensitivity and contrast, when compared to a Gaussian
beam of identical transverse resolution and optical power. This is because the Bessel beam
does not reconstruct its amplitude profile any better than the Gaussian beam when
propagating through a turbid medium that exhibits distributed scattering, as is typical with
soft tissue. We also show, however, that for fixed focus beams, Bessel beams result in
contrast and resolution superior to that of Gaussian beams, at depths where, in free-space,
they possess a higher irradiance than the Gaussian beam. To improve OCT contrast
further, alternative adaptive approaches or dynamic focussing is required.
We then demonstrate an extended-focus optical coherence microscope and use it to
perform ultrahigh-resolution optical coherence elastography, which achieves an
unprecedented (𝑥𝑥 𝑦𝑦 𝑧𝑧) 2×2×15 μm strain resolution over a depth of field of nearly 100 μm.
This is obtained by combining Bessel beam illumination and Gaussian beam detection,
compromising between depth of field improvement and contrast reduction. Our
multiphysics phase-sensitive compression OCE simulation of the influence of the system
resolution and the applied load on the measured strain precision guided our acquisition
protocols to maximise the strain sensitivity. We demonstrate this record performance on a
structured phantom and freshly excised mouse aorta.
The tools developed for image quality analysis, the characterisation of the influence of
Bessel beams on contrast in OCT, and the resolution improvement demonstrated in
ultrahigh-resolution OCE, provide important contributions to the ability of OCT and
OCT-based techniques to provide better microscopic structural and functional information
on biological tissue.
iv
Contents
Abstract ii
Contents iv
List of figures viii
List of tables xiv
Acknowledgements xv
Statement of contribution xvii
List of publications xxiii
List of acronyms xxvi
Chapter 1 Introduction 1
1.1 Research objectives .................................................................................................. 4
1.2 Structure of the thesis .............................................................................................. 4
Chapter 2 Background of OCT and OCE 9
2.1 Optical coherence tomography .............................................................................. 9
2.1.1 Low coherence interferometry ....................................................................... 16
2.1.2 Time-domain optical coherence tomography .............................................. 21
2.1.3 Fourier-domain optical coherence tomography .......................................... 26
2.2 Optical coherence elastography ............................................................................ 29
2.2.1 Elasticity ............................................................................................................ 30
2.2.2 Principles of optical coherence elastography ............................................... 35
2.3 Image quality ........................................................................................................... 38
2.3.1 OCT resolution and depth of field ................................................................ 39
2.3.2 OCT signal-to-noise ratio, sensitivity and contrast ..................................... 43
2.3.3 Speckle ............................................................................................................... 45
2.3.4 Elastogram quality............................................................................................ 47
2.4 Conclusion ............................................................................................................... 49
Chapter 3 Energy-efficient Bessel beams: a tool to alter image quality 50
3.1 Optical beam shaping ............................................................................................ 50
3.1.1 Hermite-Gaussian beams, Laguerre-Gaussian beams and non-diffracting
beams ................................................................................................................. 52
3.1.2 Beam shaping with a passive component versus an active (reconfigurable)
optical element .................................................................................................. 55
3.1.3 Location of the beam shaper in the optical path ......................................... 57
v
3.1.4 Aberration characteristics with refractive versus diffractive optical
elements .............................................................................................................59
3.2 Energy-efficient low-Fresnel-number Bessel beams and their application in optical
coherence tomography [27] ...................................................................................64
3.3 Selected dual beam OCT setup .............................................................................71
3.4 Conclusion ...............................................................................................................72
Chapter 4 Structured phantoms: a tool to mimic turbid tissue and compare image
quality 75
4.1 Introduction .............................................................................................................75
4.2 Light-tissue interaction ...........................................................................................76
4.2.1 Optical properties .............................................................................................78
4.3 Controlling the optical properties of OCT phantoms .......................................85
4.3.1 OCT silicone phantoms with varying attenuation coefficients .................86
4.3.2 OCT silicone phantoms with varying anisotropy ........................................87
4.3.3 Scattering overlayers validation ......................................................................88
4.4 Mechanical properties.............................................................................................90
4.4.1 Controlling the mechanical properties of OCT silicone phantoms ..........92
4.5 Structured three-dimensional optical phantom for optical coherence tomography
[30] .............................................................................................................................95
4.5.1 Introduction ......................................................................................................95
4.5.2 Method ...............................................................................................................96
4.5.3 Results and discussion .....................................................................................99
4.5.4 Conclusions .................................................................................................... 102
4.5.5 Aknowledgements ......................................................................................... 102
4.6 Image quality test targets ..................................................................................... 103
4.6.1 Point spread function and contrast phantoms characteristics ................ 103
4.6.2 Phantom validation ....................................................................................... 105
4.7 Elastography structured phantoms.................................................................... 108
4.8 Conclusion ............................................................................................................ 110
Chapter 5 Simulation of beam propagation and image formation in turbid samples:
a tool to theoretically quantify image quality 111
5.1 Introduction .......................................................................................................... 111
5.2 OCT image simulation in the single-scattering regime ................................... 112
5.2.1 OCT image simulation as local sums of random phasors ....................... 115
5.2.2 OCT image simulation as superposition of linear system responses ..... 120
vi
5.3 A full wave 2-D model of image formation in optical coherence tomography
applicable to general samples [32] ................................................................... 125
5.3.1 Details of the model ...................................................................................... 128
5.3.2 Evaluation and analysis ................................................................................. 133
5.3.3 Discussion and conclusions .......................................................................... 144
5.3.4 Acknowledgments .......................................................................................... 144
5.4 3-D simulation of optical beam propagation in phantoms ............................ 145
5.4.1 Simulation of beam propagation in free-space .......................................... 145
5.4.2 Simulation of beam propagation in tissue phantoms ............................... 147
5.5 Conclusion ............................................................................................................. 149
Chapter 6 Quantifying OCT image quality in turbid samples using Gaussian and
Bessel beams 151
6.1 Introduction .......................................................................................................... 151
6.1.1 Testing arrangement: sample configuration with a Gaussian beam ....... 152
6.2 Quantifying the influence of Bessel beams on image quality in optical coherence
tomography [28].................................................................................................... 155
6.2.1 Introduction .................................................................................................... 155
6.2.2 Results .............................................................................................................. 158
6.2.3 Discussion ....................................................................................................... 164
6.2.4 Methods ........................................................................................................... 168
6.3 Conclusion ............................................................................................................. 171
Chapter 7 Improving OCE image quality: strain precision and resolution 173
7.1 Introduction .......................................................................................................... 173
7.1.1 Phase-sensitive OCT displacement measurement precision ................... 173
7.2 Analysis of image formation in optical coherence elastography using a multiphysics
approach [25] ......................................................................................................... 177
7.2.1 Introduction .................................................................................................... 177
7.2.2 Metrics of elastogram quality and precision ............................................... 179
7.2.3 Multiphysics model of optical coherence elastography ............................ 180
7.2.4 Experimental procedure ................................................................................ 184
7.2.5 Results .............................................................................................................. 187
7.2.6 Discussion ....................................................................................................... 190
7.2.7 Implications for the ultrahigh-resolution regime ...................................... 194
7.3 Ultrahigh-resolution optical coherence elastography [31] ............................. 196
7.4 Conclusion ............................................................................................................. 203
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Chapter 8 Conclusions 205
8.1 Research contributions and significance ........................................................... 206
8.2 Study limitations and future work ..................................................................... 211
8.2.1 Current limitations ......................................................................................... 211
8.2.2 Proposed future work ................................................................................... 213
8.3 Final remarks ........................................................................................................ 214
Appendix A Spatial light modulator characterisation 215
Bibliography 225
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List of figures
Figure 2-1. Schematic of terminology used when referring to OCT scan orientation.
......................................................................................................................... 10
Figure 2-2. Spectral absorption for a range of tissue chromophores in the diagnostic
window in the near-infrared. ....................................................................... 11
Figure 2-3. Comparison of OCT resolution and imaging depths to other biomedical
imaging methods. .......................................................................................... 12
Figure 2-4. Bar chart of a snapshot of results from a Scopus search of selected OCT
biological applications from 2005 to 2015, inclusive. .............................. 13
Figure 2-5. Main applications of OCT. ............................................................................. 13
Figure 2-6. Video showcasing representative examples of the main clinical and
laboratory research applications of OCT. ................................................. 14
Figure 2-7. Video showcasing areas of fundamental research aimed at image quality
improvement in OCT. .................................................................................. 14
Figure 2-8. Optical coherence tomography (OCT) setups. ............................................ 15
Figure 2-9. Transform-limited pulse generation by addition of weighted spectral
components with their phases locked to each other. ............................... 17
Figure 2-10. Temporal evolution of polychromatic wave generated by addition of
weighted spectral components with their phases randomly assigned. .. 18
Figure 2-11. Temporal evolution of monochromatic (left) wave and a polychromatic
CW (right) wave. ........................................................................................... 19
Figure 2-12. Spectra (in green) of the monochromatic (left) and polychromatic (right)
waves shown in Figure 2-11 after 143 ps. ................................................. 19
Figure 2-13. Autocorrelation of the monochromatic (left) and polychromatic (right)
waves shown in Figure 2-12. ....................................................................... 20
Figure 2-14. Equivalence of temporal autocorrelation of polychromatic waves
generated by a transform-limited pulse (left) and by a continuous wave
(CW) (right). ................................................................................................... 21
Figure 2-15. Interferometry with a monochromatic wave. ............................................. 23
Figure 2-16. TD-OCT photocurrent over time. .............................................................. 24
Figure 2-17. Field amplitudes (left panel), electric charge generated at the
photodetector (middle panel) and detector photocurrent as a function
of reference arm group delay. ...................................................................... 25
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Figure 2-18. An example of the spectral interference generated by a discrete sample
reflectivity function (top), and the corresponding A-scan captured by
FD-OCT (bottom). .......................................................................................28
Figure 2-19. Reported values and ranges of Young’s modulus for selected tissues and
tissue constituents. .........................................................................................34
Figure 2-20. Illustration of phase-sensitive compression optical coherence
elastography on a structured phantom. ......................................................37
Figure 2-21. Optical coherence elastography of a malignant breast tumour with
quantitative elasticity estimation. .................................................................38
Figure 2-22. Schematic illustrating diffraction-limited resolution in microscopy. .......40
Figure 2-23. Schematic illustrating the determinants of resolution in OCT. ...............42
Figure 2-24. Calculated sensitivity for a typical spectrometer-based SD-OCT system.
..........................................................................................................................44
Figure 2-25. OCT image of a human fingertip with high contrast speckle pattern
plotted on a logarithmic grayscale. ..............................................................45
Figure 2-26. Marginal probability density function 𝑝𝑝𝑝𝑝𝑝𝑝 for the OCT signal phase. .48
Figure 3-1. Trade-off between the DOF and transverse resolution in OCT, at
wavelength of 0.84 µm in air. ......................................................................51
Figure 3-2. Collimated Gaussian beam shaped and focussed by different lenses. ......54
Figure 3-3. Examples of shaped beams. ............................................................................55
Figure 3-4. Static optical elements: lenses. .........................................................................56
Figure 3-5. Reconfigurable beam shapers. .........................................................................56
Figure 3-6. Bessel beam and its typical spatial frequency spectrum. .............................58
Figure 3-7. Typical experimental configurations for the generation of Bessel beams.
..........................................................................................................................58
Figure 3-8. Refractive and diffractive optical elements. ..................................................60
Figure 3-9. Simulations of the beam profile of a Bessel beam, using VirtualLab. .......61
Figure 3-10. Graphical illustration of the cause of the achromatic characteristics of a
DOE-generated Bessel beam. ......................................................................61
Figure 3-11. Simulation results for the diffraction efficiency of an SLM-generated
Bessel beam. ...................................................................................................62
Figure 3-12. Energy efficiency of a Bessel beam compared to a Gaussian beam of
same NA. .........................................................................................................66
Figure 3-13. Experimental setup for OCT imaging using low-Fresnel-number Bessel
beams. ..............................................................................................................68
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Figure 3-14. Experimental results comparing the imaging performance of low-
Fresnel-number Bessel beams and a Gaussian beam of the same
transverse resolution. .................................................................................... 69
Figure 3-15. Dual-beam OCT system schematic. ............................................................ 72
Figure 3-16. Self reconstruction property of a Bessel beam........................................... 73
Figure 4-1. Scattering interaction in tissue classified into three categories. ................. 78
Figure 4-2. Mie theory of scattering of a plane wave by a sperical dielectric particle. 81
Figure 4-3. Typical average OCT signal, on a logarithmic scale, as a function of
depth. .............................................................................................................. 83
Figure 4-4. OCT signal characteristics as a function of increasing scatterer
concentration. ................................................................................................ 86
Figure 4-5. Scattering overlayers. ........................................................................................ 88
Figure 4-6. Measurements of elasticity and viscoelasticity of materials and apparatus.
......................................................................................................................... 92
Figure 4-7. Comparison of range of Young’s moduli of phantom materials and soft
tissue. ............................................................................................................... 93
Figure 4-8. 3-D structured phantom design and characterisation. ................................ 97
Figure 4-9. OCT images of Phantom I and II. ............................................................... 100
Figure 4-10. Video of sequential multiplanar views and a fly-through of the 3-D solid
rendering of the phantom. ......................................................................... 100
Figure 4-11. Speckle reduction performed on Phantom II. ......................................... 101
Figure 4-12. Imaging targets. ............................................................................................. 104
Figure 4-13. OCT characterisation of the imaging targets with Gaussian and Bessel
beams. ........................................................................................................... 106
Figure 4-14. Ultrahigh-resolution OCE inclusion phantom. ....................................... 109
Figure 5-1. Simulation of the axial distortion of an OCT A-scan caused by speckle as
a result of the coherent detection process. .............................................. 113
Figure 5-2. Simulation of the one-dimensional convolution operation resulting in the
single-scattering OCT A-scan signal phasor. .......................................... 114
Figure 5-3. Comparison between images of a structured phantom acquired using a
coherent (OCT) system (a) and a bright field microscope, i.e., an
incoherent system (b). ................................................................................ 119
Figure 5-4. Simulation of (a) a hypothetical incoherent and (b) a coherent (OCT)
image formation process, according to the simple model presented in
this section.................................................................................................... 120
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Figure 5-5. Illustration of the model of image formation in OCT, and comparison
with experimental results. .......................................................................... 123
Figure 5-6. A schematic of the modelled optical system. ............................................. 129
Figure 5-7. Schematic diagram of where the source field is introduced 𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆 and
where the scattered field is recorded 𝑆𝑆𝑆𝑆𝑆𝑆 in FDTD simulations. ........ 132
Figure 5-8. Numerical dispersion. .................................................................................... 135
Figure 5-9. Simulation of the OCT image of a stratified medium. ............................. 136
Figure 5-10. Images and plots which demonstrate the depth-dependent PSFs
employed in OCT by simulating images of 24 scatterers arranged
equidistantly along the optical axis. .......................................................... 137
Figure 5-11. 3-D structured phantom for OCT. ........................................................... 140
Figure 5-12. Diagram showing how a collection of spherical scatterers in a three-
dimensional beam (left) can be approximated in a two-dimensional
system (right). .............................................................................................. 140
Figure 5-13. Simulation of OCT image formation for a structured phantom. ......... 141
Figure 5-14. Optical beams: Gaussian and Bessel beams of equal resolution and
power. ........................................................................................................... 146
Figure 5-15. Illustration of the EM simulation. ............................................................. 147
Figure 5-16. (a) Simulated and (b) experimental transverse PSFs for Gaussian and
Bessel beams, top and bottom, respectively. .......................................... 149
Figure 6-1. Effect of increasing degree of wide-angle multiple scattering
(Category III) signal on OCT images of the two different structured
phantoms, described in Section 4.5. ........................................................ 153
Figure 6-2. Effect of increasing degree of low-angle forward scattering (Category II)
signal on OCT images of the Structured Phantom I overlaid by
phantoms containing different diameter polystyrene beads. ................ 154
Figure 6-3. Beam and sample configuration. (a) Schematic diagram of the beam,
overlayer and imaging target, for transparent (top) and scattering
overlayers (bottom). ................................................................................... 157
Figure 6-4. Simulated beam profiles after propagation through Overlayers 1–3 with
increasing anisotropy from left to right compared to (left) propagation
through a scattering-free (SF) overlayer. ................................................. 159
Figure 6-5. Simulated transverse beam profiles. ............................................................ 160
Figure 6-6. Simulated total beam irradiance and image-carrying and -degrading
components of the beam versus depth in the PSF phantom after each
overlayer. ...................................................................................................... 161
xii
Figure 6-7. On-axis signal-to-background ratio (SBR) versus depth for Overlayers 1–
3. .................................................................................................................... 162
Figure 6-8. OCT contrast assessment. ............................................................................. 163
Figure 6-9. Angular spectra intensity of the beams. ...................................................... 166
Figure 7-1. Origin of translation-induced phase decorrelation in one dimension. ... 175
Figure 7-2. Origin of strain-induced phase decorrelation in one dimension. ............ 176
Figure 7-3. FEM simulation of a sample containing a stiff inclusion under quasi-static
compression. ................................................................................................ 181
Figure 7-4. Computing the new location of the scattering potentials under an applied
load. ............................................................................................................... 183
Figure 7-5. Flowchart of the multiphysics simulation of OCE. .................................. 183
Figure 7-6. Inclusion phantom and phase-sensitive compression OCE signal
signatures. ..................................................................................................... 186
Figure 7-7. Experimental scans of a silicone inclusion phantom compared to results
of the multiphysics simulation of phase-sensitive compression OCE.
....................................................................................................................... 187
Figure 7-8. Regions used for comparing experiment to simulation, shown on (a) the
simulated OCT image, and (b) the simulated strain elastogram. ......... 188
Figure 7-9. Displacement sensitivity (𝑆𝑆𝑠𝑠) vs. (a) local strain at various depths in the
sample, and (b) vs. depth at selected values of strain in the sample. ... 189
Figure 7-10. Strain SNR (SNR𝜀𝜀) (a) vs. local strain at various depths in the sample,
and (b) vs. depth at selected values of strain in the sample. ................. 190
Figure 7-11. Effect of the OCT axial resolution on the precision of the sample axial
displacement measurement. ....................................................................... 195
Figure 7-12. UHROCE system. ........................................................................................ 197
Figure 7-13. OCT images and strain elastograms of an inclusion phantom taken with
two systems. ................................................................................................. 200
Figure 7-14. OCT images and strain elastograms of a mouse aorta taken with the two
systems: OCE and UHROCE, compared with histology. .................... 202
Figure A-1. Illustration of the connection diagram of the SLM, with a picture of one
module, a close-up of the LCoS screen, with a schematic of the pixel
structure and the operating principle. ...................................................... 216
Figure A-2. Installed and connected SLM. ..................................................................... 217
Figure A-3. Testing setup for the SLM characterisation. .............................................. 217
Figure A-4. Setup to test the effect of phase droop on the Gaussian beam focus. .. 219
xiii
Figure A-5. Time sequence over 8.34 ms of 8 images of the SLM screen and
corresponding transverse focal intensity. ................................................ 219
Figure A-6. Recurrent defocus. ........................................................................................ 220
Figure A-7. Time-resolved (over 50 ms) powers in the 0th and 1st diffractive orders as
a function of the modulation depth discretization and grating period for
the two SLM modules loaded with different configurations. .............. 221
Figure A-8. Phase droop as a function of the corresponding grey level for the two
SLM modules and various configurations. ............................................. 222
Figure A-9. SLM phase calibration to obtain a linear conversion from grey scale to
phase retardation. A 256 grey level offset corresponds to a 2π phase
shift for a given wavelength. ..................................................................... 223
Figure A-10. Characterisation of the linearity of the phase retardation with grey level.
....................................................................................................................... 224
xiv
List of tables
Table 2-1. Imaging parameters for OCE displacement estimation methods. Adapted
from [18]. ........................................................................................................ 35
Table 2-2. Experimental and theoretical imaging parameters for OCE loading
methods. Adapted from [18]. ...................................................................... 35
Table 3-1. Experimental comparison Bessel vs. Gaussian ............................................. 70
Table 4-1. Specified and calculated scattering overlayer characteristics ....................... 87
Table 4-2. Young’s modulus of various mixing ratios (Cross-linker: Catalyst) for
Wacker Elastosil 601 silicone phantoms ................................................... 92
Table 4-3. Young’s modulus of various mixing ratios (Cross-linker: Catalyst: PDMS
oil) for Wacker Elastosil 601 silicone and Wacker AK50 PDMS oil
phantoms ........................................................................................................ 93
Table 4-4. Contrast phantom nominal characteristics ................................................... 105
Table 5-1. Simulated and experimental beam characteristics ....................................... 145
Table 5-2. Simulation characteristics ................................................................................ 148
Table 6-1. Reduction in contrast ...................................................................................... 163
Table 6-2. Simulated and experimental beam characteristics ....................................... 169
Table 6-3. Specified and calculated scattering overlayer characteristics ..................... 169
Table 6-4. Contrast phantom nominal characteristics ................................................... 170
Table 7-1. OCE simulation inputs and parameters ....................................................... 184
Table 7-2. Numerical comparison of experimentally acquired vs. simulated
elastograms for the regions marked in Figure 7-8. ................................. 188
xv
Acknowledgements
Many people have helped, directly or indirectly, to shape this thesis, and I personally want
to acknowledge them for their contribution and support.
First, David Sampson, for giving me the opportunity to join the Optical + Biomedical
Engineering Laboratory (OBEL) in 2008 and, after several years of research appointments,
encouraging me to pursue a PhD degree to capitalise on my experience. Thanks David for
mentoring me, providing and leading an excellent research facility, with high-profile
international collaborations and a very healthy work environment that promotes work-life
balance. I also feel very grateful to my other supervisors, Peter Munro and Dirk Lorenser,
who have taught me a great deal of knowledge and helped me manage this research project
effectively, navigating through a lot of unknowns and difficulties, technical or otherwise. I
am also very thankful to The University of Western Australia for supporting me through an
Australian Postgraduate Award scholarship.
Special thanks goes to my newfound home of Western Australia and Perth. It helped
me connect my work with the places and nature around me through countless and
beautiful examples of the ubiquity of the topics being studied in this thesis. These awe-
inspiring optical and physical phenomena might take a different form from those
researched, but they are all related: light scattering by the west coast pristine sky in
enchanting sunsets, refraction by water droplets in Perth’s winter rainbows, reduced
visibility by stormy and foggy clouds over the river, wave interference patterns in
Freshwater bay, summer light attenuation by sunscreen at Floreat Beach.
My professional development and research creativity have greatly benefitted from in-
depth discussions and guidance by colleagues and high-calibre researchers Timothy
Hillman, Brendan Kennedy, Maciej Wojtkowski and Martin Villiger. Without their input
and our exchange of ideas, the richness of this research would have suffered.
I want to thank Blake Klyen and Loretta Scolaro for having been, not only peers
providing the biggest continuing support and having shared their research experience with
me, but also for being true friends.
Other colleagues and fellow PhD candidates have been a source of knowledge and
inspiration throughout the duration of this research: Bryden with his fearless “can do”
attitude, Philip with his humbleness and wit, Kelsey with her positive work ethics, Lixin
with his competence and diligence, Robert with his focus and drive and the rest of the
OBEL team and collaborators with their unique characteristics.
xvi
I cannot forget the moral support and keen interest from Cibele in my research and my
career development, which strengthened my confidence in the pursuit of this achievement.
Roberta, Laurent, Monica, Seon, Danila, Uros, Cris, Valerio and Jana, you are all good
friends and I am fortunate to have you in my life, and I am sure you are all delighted by this
accomplishment. Through your smiles, our laughs and the good times together, you have
helped me strike the right balance, and tackle the intricacies of research with a positive
attitude. Thanks to my housemates and friends, especially Kat, Lana, Dario and Jon who
have been supportive and understanding at all times.
Thanks to all the UWA OSA student chapter members, and Danka in particular, for
having created something fun and valuable in Perth for the young and upcoming optics
and photonics community, through professional development and scientific outreach
efforts.
Mens sana in corpore sano. A very true Latin statement, and for me a healthy body comes
from the sporting activity that I enjoy the most: volleyball. So thanks to all my volleyball
and beach volleyball mates.
Heather and Chris, you have been like family to me here in Perth. From the very
beginning, you helped me settle in the very best of ways, and I will always be grateful to
you for that and the warmth you gave me.
Last, but not least, thanks to my friends and family back home in Italy. Thanks to
Francesca, Chino and Andrea. Your affection has travelled through continents. Grazie to
my close relatives, my cousins, my late grandparents, my brother Marco and my parents
Lucia and Roberto. Your unconditional encouragement and love has provided, at the same
time, a safe haven to reflect upon my values and interests, and a springboard that
strengthened my motivation to look ahead and to reach new grounds.
xvii
Statement of contribution
This thesis contains the results of the research that I, Andrea Curatolo (AC), performed
within the School of Electrical, Electronic, and Computer Engineering at The University of
Western Australia, between 2011 and 2016. Except where indicated below and throughout
the text, all work and writing are my own. The chapters of this thesis are primarily derived
from nine published works: four first-authored, journal articles (three full-length and one
letter); three co-authored, journal articles (two full-length and one letter); one co-authored
review article; and one first-authored book chapter1. I am the sole author of the remainder
of the document. Sections 3.2, 4.5, 5.3, 6.2, and 7.3 are reproductions of publications, as
reported below, included without change in content, but modified only in formatting for
consistency with the remainder of this document. I am the sole author of Chapters 1, 8,
Appendix A and the vast majority of Chapter 2.
Listed below are the contributions of each author to each publication. My contributions
are given as a percentage next to my name, and in the descriptions.
1. B. F. Kennedy, R. A. McLaughlin, K. M. Kennedy, L. Chin, A. Curatolo
(15%), A. Tien, B. Latham, C. M. Saunders, and D. D. Sampson, "Optical
coherence micro-elastography: mechanical-contrast imaging of tissue
microstructure," Biomedical Optics Express 5, 2113-2124 (2014).
Publication 1 features in parts in Section 2.2 and in Section 4.7.
B. F. Kennedy (BFK) was the principle author of this article. BFK led the design of the
system, and all experiments, assisted with the design of the signal processing code, and led
the writing of the manuscript, which was reviewed and edited by all co-authors. R. A.
McLaughlin (RAM) assisted with analysis of results, provided software for, and assistance
with, co-registration with histology, organised access to tissue and clinical collaboration,
and helped develop the signal processing code. K. M. Kennedy (KMK) fabricated
phantoms, helped performed phantom imaging, tissue imaging, and interpretation of
results. L. Chin (LC) helped develop the signal processing chain, performed data
processing, helped perform tissue imaging and generated schematic figures. AC assisted
with early technology development, phantom and tissue imaging, and generated schematic
figures. A. Tien (AT) obtained patient consent, facilitated access to tissue, and in some
1 This thesis was prepared according to The University of Western Australia’s guidelines on “Thesis ss a series of papers”
(http://www.postgraduate.uwa.edu.au/students/thesis/series. Accessed 27 June 2016).
xviii
cases performed surgery. B. Latham (BL) interpreted histopathology for diagnosis, and
assisted co-registration to OCT and OCE data. C. M. Saunders (CMS) performed surgeries,
aided in the conceptual development of the research, and facilitated access to tissue. D. D.
Sampson (DDS) conceptualised the work, provided overall guidance, facilitated
collaboration, and supervised drafting of the manuscript.
2. L. Chin, A. Curatolo (40%), B. F. Kennedy, B. J. Doyle, P. R. T. Munro, R. A.
McLaughlin, and D. D. Sampson, “Analysis of image formation in optical
coherence elastography using a multiphysics approach,” Biomedical Optics
Express 5, 2913–2930 (2014).
Publication 2 features in parts in Section 2.2.2, in Section 4.7 and in Section 5.2.2. Section
7.2 includes it from the methods section until the conclusion.
LC and AC were the joint principle authors of this article. AC fabricated the inclusion
and homogeneous phantoms, performed the phantom imaging experiments, led the design
of the optical simulation, performed the OCE simulation of the homogeneous phantoms,
led the data analysis and generated the first draft of the figures. LC led the design of the
mechanical simulation, and the combination of the optical and mechanical models,
optimised the simulation code, performed the OCE simulation of the inclusion phantom,
refined the figures, and led the writing of the manuscript, which was reviewed and edited
by all co-authors. BFK provided guidance and supervision, and facilitated collaboration. B.
J. Doyle (BJD) provided advice and training on the mechanical simulation. P. R. T. Munro
(PRTM) provided advice and guidance on the optical simulation. RAM provided advice on
the simulation code. DDS provided overall guidance and supervision of the research.
3. A. Curatolo (60%), B. F. Kennedy, D. D. Sampson, and T. R. Hillman,
"Speckle in Optical Coherence Tomography," in Advanced Biophotonics: Tissue
Optical Sectioning (V. V. Tuchin, and R. Wang, eds), Taylor & Francis: 211-277
(2013).
Publication 3 features in parts in Section 2.3.3, in Section 4.2 and in Section 5.2.1.
AC was the principle author of this book chapter. AC prepared an extensive literature
review, proposed the chapter structure, organised the material, collected and updated
existing figures, generated new figures, and led the writing of the chapter, which was
reviewed and edited by all co-authors. BFK led the writing of Section 6.7 about mitigation
of OCT speckle. DDS conceptualised the work, provided overall guidance, facilitated the
discussion and timeline management with the editors, and supervised drafting of the
xix
manuscript. T. R. Hillman (TRH) provided published and unpublished material for the
chapter and strong technical and writing guidance.
4. D. Lorenser, C. C. Singe, A. Curatolo (20%), and D. D. Sampson, "Energy-
efficient low-Fresnel-number Bessel beams and their application in optical
coherence tomography," Optics Letters 39, 548-551 (2014).
Publication 4 appears in full as published in Section 3.2.
D. Lorenser (DL) was the principle author of this article. DL led the engineering and
development of the theory of low-Fresnel Bessel beams and their energy efficiency in
comparison to equal resolution Gaussian beams. DL designed the dual beam OCT system
and the software to extract the DOF improvement and OCT sensitivity penalty from OCT
images of a nanoparticle embedded phantom. He also led the writing of the manuscript,
which was reviewed and edited by all co-authors. C. C. Singe (CCS) contributed to the
system design through simulations, built parts of the OCT system, aligned it and help
characterise it. AC chose, characterised and calibrated the beam shaper. He built parts of
the OCT system, measured and verified its specifications. He also acquired and processed
the images of phantoms and biological tissue, and helped prepare some of the manuscript
figures. DDS conceptualised the work, provided overall guidance and supervision of the
research.
5. A. Curatolo (65%), P. R. T. Munro, D. Lorenser, P. Sreekumar, C. C. Singe, B.
F. Kennedy, and D. D. Sampson, "Quantifying the influence of Bessel beams
on image quality in optical coherence tomography," Scientific Reports 6, 23483
(2016).
Publication 5 features in parts of its supplementary information in Section 3.3, in Section
4.3, in Section 4.6 and in Section 5.4. The main manuscript also appears in full as published
in Section 6.2.
AC was the principle author of this article. AC prepared an extensive literature review
and proposed the image quality benchmarking setup. AC also rebuilt, aligned and
characterised the selected dual beam OCT setup. He designed and validated the scattering
overlayers, i.e., the tissue-mimicking phantoms with varying scattering anisotropy. He
designed and sourced the 3-D structured phantom used as a contrast target. AC wrote code
to process the OCT images, to verify phantom specifications, to model the numerical
phantoms and to measure and analyse OCT contrast. AC performed the experiments and
processed the simulation results. AC generated the manuscript figures and led the writing
xx
of the article, which was reviewed and edited by all co-authors. PRTM designed, provided
and run the simulation code to analyse the beam propagation through the scattering
numerical phantoms using a rigorous full-wave solver of Maxwell’s equations. He also
provided supervision of the project, manuscript overview, guidance and insight into
scattering phenomena interpretation. DL supervised the dual beam OCT setup redesign
and characterisation, provided supervision of the project, and useful discussion about the
validation of the imaging test target. P. Sreekumar (PS) manufactured the scattering
overlayers and helped in their validation and verification of their specifications. CCS helped
in the alignment and characterisation of the dual beam OCT setup and in the initial stages
of the scattering overlayers validation. BFK provided help and insight in the design of the
contrast target. DDS conceptualised the work, provided overall guidance and supervision
of the research with insightful discussions.
6. G. Lamouche, B. F. Kennedy, K. M. Kennedy, C.-E. Bisaillon, A. Curatolo
(10%), G. Campbell, V. Pazos, and D. D. Sampson, "Review of tissue
simulating phantoms with controllable optical, mechanical and structural
properties for use in optical coherence tomography," Biomedical Optics
Express 3, 1381-1398 (2012).
Publication 6 features in parts from Section 4.1 to Section 4.4.
Guy Lamouche (GL) and BFK were the joint principle authors of this article. GL led
the writing of the manuscript, which was reviewed and edited by all co-authors. BFK
supervised the writing of all sections relating to mechanical properties, fibrin phantoms,
and complex structures. KMK led the writing of the mechanical properties section of the
article. C.-E. Bisaillon (CEB) led the experimental work on PVA phantoms and
characterisation of optical properties. AC led the writing of the first part of the complex
structures section of the article. G. Campbell (GC) consulted on the artery phantom work.
V. Pazos (VP) consulted on sections regarding PVA phantoms. DDS supervised the
drafting of the manuscript.
7. A. Curatolo (70%), B. F. Kennedy, and D. D. Sampson, "Structured three
dimensional optical phantom for optical coherence tomography," Optics
Express 19, 19480-19485 (2011).
Publication 7 appears in full as published in Section 4.5.
AC was the principle author of this article. AC contracted the Australian National
Fabrication Facility (ANFF) node with equipment and expertise in advanced micro-fluidic
xxi
device fabrication, with the goal of manufacturing a structured OCT phantom. AC
designed the 3-D structured phantom, and pursued its validation through several
interactions with the ANFF by overcoming manufacturing constraints. He then verified the
specifications of the four different manufactured types. He performed the experiments and
the image and data processing. He also generated the manuscript figures and led the writing
of the article, which was reviewed and edited by all co-authors. BFK supervised and helped
in the discussions with the ANFF and provided valuable feedback on the design
adjustments to suit manufacturing constraints. DDS advocated and inspired the start of
this research and provided overall guidance and supervision.
8. P. R. T. Munro, A. Curatolo (15%), and D. D. Sampson, "Full wave model of
image formation in optical coherence tomography applicable to general
samples," Optics Express 23, 2541-2556 (2015).
Publication 8 appears in full as published in Section 5.3.
PRTM was the principle author of this article. PRTM conceptualised this research,
designed the OCT image simulator and its various modules, including the rigorous
full-wave solver of Maxwell’s equations to calculate the spatial distribution of the scattered
light field form the sample. He wrote the code, and performed the simulations. He also
generated the majority of the manuscript figures and led the writing of the article, which
was reviewed and edited by all co-authors. AC provided feedback on the way to implement
the OCT working principles in the simulation. AC performed the experiments with the
structured phantom, processed the images and helped in generating the related figure. He
also provided information for the input simulation parameters and feedback on the
simulation results verification. DDS conceptualised the work, provided overall guidance
and supervision.
9. A. Curatolo (50%), M. Villiger, D. Lorenser, P. Wijesinghe, A. Fritz, B. F.
Kennedy, and D. D. Sampson, "Ultrahigh-resolution optical coherence
elastography," Optics Letters 41, 21-24 (2016).
Publication 9 appears in full as published in Section 7.3.
AC was the principle author of this article. AC contributed to the hardware design of
the extended-focus optical coherence microscope (xf-FDOCM) and to building it. He
aligned and characterised the system. He devised acquisition strategies to acquire 3-D
elastograms with it and to process them. He designed and manufactured suitable inclusion
phantoms. He performed the experiments and processed the images and data. He liaised
xxii
with CellCentral and provided the tissue for histology. He also generated the manuscript
figures and led the writing of the article, which was reviewed and edited by all co-authors.
M. Villiger (MV) participated in the project conceptualisation. He also contributed to the
hardware design, and to its characterisation. He helped with the spectrometer calibration
and provided useful feedback throughout the project duration. DL participated in the
project conceptualisation. He designed and chose the components for the xf-FDOCM
system. He closely supervised the building of it. PW provided support in the discussions
about the acquisition strategies. He also refined the image reconstruction and processing
code. He helped in the image acquisition effort and some of the experiments. He also
scanned the histology sections, which formed part of Figure 3. A. Fritz (AF) helped in
building and aligning the xf-FDOCM system. BFK participated in the project
conceptualisation and provided close supervision throughout the whole project. He gave
insightful feedback on all elastography aspects of the research and sourced the mouse aorta
for tissue imaging. DDS conceptualised the work, and provided overall guidance and
supervision.
xxiii
List of publications
The following is a chronological list of publications arising during the duration of this
thesis. PDFs and online multimedia material of some of these publications are available on
the website of the Optical + Biomedical Engineering Lab (OBEL):
http://obel.ee.uwa.edu.au/publications.
Fully refereed journal articles
Key: † These authors contributed equally to this work
1. A. Curatolo, B. F. Kennedy, and D. D. Sampson, "Structured three dimensional
optical phantom for optical coherence tomography," Optics Express 19, 19480-
19485 (2011).
2. R. A. McLaughlin, B. C. Quirk, A. Curatolo, R. W. Kirk, L. Scolaro, D. Lorenser, P.
D. Robbins, B. A. Wood, C. M. Saunders, and D. D. Sampson, "Imaging of Breast
Cancer With Optical Coherence Tomography Needle Probes: Feasibility and Initial
Results," IEEE Journal of Selected Topics in Quantum Electronics 18, 1184-1191
(2012).
3. A. Curatolo†, R. A. McLaughlin†, B. C. Quirk, R. W. Kirk, A. G. Bourke, B. A.
Wood, P. D. Robbins, C. M. Saunders, and D. D. Sampson, "Ultrasound-Guided
Optical Coherence Tomography Needle Probe for the Assessment of Breast
Cancer Tumor Margins," American Journal of Roentgenology 199, W520-W522
(2012).
4. G. Lamouche, B. F. Kennedy, K. M. Kennedy, C.-E. Bisaillon, A. Curatolo, G.
Campbell, V. Pazos, and D. D. Sampson, "Review of tissue simulating phantoms
with controllable optical, mechanical and structural properties for use in optical
coherence tomography," Biomedical Optics Express 3, 1381-1398 (2012).
5. D. Lorenser, C. C. Singe, A. Curatolo, and D. D. Sampson, "Energy-efficient low-
Fresnel-number Bessel beams and their application in optical coherence
tomography," Optics Letters 39, 548-551 (2014).
6. B. F. Kennedy, R. A. McLaughlin, K. M. Kennedy, L. Chin, A. Curatolo, A. Tien,
B. Latham, C. M. Saunders, and D. D. Sampson, "Optical coherence micro-
elastography: mechanical-contrast imaging of tissue microstructure," Biomedical
Optics Express 5, 2113-2124 (2014).
xxiv
7. L. Chin†, A. Curatolo†, B. F. Kennedy, B. J. Doyle, P. R. T. Munro, R. A.
McLaughlin, and D. D. Sampson, "Analysis of image formation in optical
coherence elastography using a multiphysics approach," Biomedical Optics Express
5, 2913-2930 (2014).
8. P. R. T. Munro, A. Curatolo, and D. D. Sampson, "Full wave model of image
formation in optical coherence tomography applicable to general samples," Optics
Express 23, 2541-2556 (2015).
9. B. F. Kennedy, R. A. McLaughlin, K. M. Kennedy, L. Chin, P. Wijesinghe, A.
Curatolo, A. Tien, M. Ronald, B. Latham, C. M. Saunders, and D. D. Sampson,
"Investigation of optical coherence micro-elastography as a method to visualize
cancers in human breast tissue," Cancer Research 75, 3236-3245 (2015).
10. A. Curatolo, M. Villiger, D. Lorenser, P. Wijesinghe, A. Fritz, B. F. Kennedy, and
D. D. Sampson, "Ultrahigh-resolution optical coherence elastography," Optics
Letters 41, 21-24 (2016).
11. A. Curatolo, P. R. T. Munro, D. Lorenser, P. Sreekumar, C. C. Singe, B. F.
Kennedy, and D. D. Sampson, "Quantifying the influence of Bessel beams on
image quality in optical coherence tomography," Scientific Reports 6, 23483 (2016).
Selected conference papers
Key: * International | ^ Domestic | † Full paper| § Abstract|
1. *§ A. Curatolo, B. F. Kennedy, K. M. Kennedy, R. A. McLaughlin, and D. D.
Sampson, "Portable 3D optical coherence elastography for applying microscopic
imaging of tissue mechanical properties in the clinic," in Optics Within Life Science
(Genoa, Italy, 2012).
2. *§ A. Curatolo, L. Chin, C. Stynes, B. F. Kennedy, and D. D. Sampson, "Simulation
of image formation process in phase-sensitive optical coherence elastography," in
Opto, Meeting for Young Researchers (Torun, Poland, 2013).
3. ^§ A. Curatolo, C. C. Singe, D. Lorenser, P. R. T. Munro, and D. D. Sampson,
"Improving OCT image quality in turbid structured phantoms by beam shaping," in
ANZCOP (Fremantle, Australia, 2013).
4. *§ A. Curatolo, D. Lorenser, P. R. T. Munro, P. Sreekumar, C. C. Singe, B. F.
Kennedy, and D. D. Sampson, "Analysis of beam shaping in optical coherence
tomography in the presence of sample-induced aberrations and scattering," in SPIE
Photonics West, BiOS: Optical Coherence Tomography and Coherence Domain Optical Methods
in Biomedicine XIX (San Francisco, USA, 2015).
xxv
5. *§ A. Curatolo, M. L. Villiger, D. Lorenser, A. Fritz, B. F. Kennedy, and D. D.
Sampson, "Cellular resolution optical elastography using phase-sensitive extended
depth-of-field optical coherence microscopy," in SPIE Photonics West, BiOS: Optical
Elastography and Tissue Biomechanics II (San Francisco, USA, 2015).
6. *§ A. Curatolo, P. R. T. Munro, P. Sreekumar, C. C. Singe, B. F. Kennedy, D.
Lorenser, and D. D. Sampson, "Effect on optical coherence tomography image
quality of turbid tissue scattering using Gaussian or Bessel beams," in SPIE ECBO
2015, European Conference in Biomedical Optics (Munich, Germany, 2015).
7. *† A. Curatolo, M. L. Villiger, D. Lorenser, A. Fritz, B. F. Kennedy, and D. D.
Sampson, "Ultrahigh-resolution optical coherence elastography using a Bessel beam
for extended depth of field," in SPIE Photonics West, BiOS: Optical Coherence
Tomography and Coherence Domain Optical Methods in Biomedicine XX (San Francisco,
USA, 2016).
Book chapters
A. Curatolo, B. F. Kennedy, D. D. Sampson, and T. R. Hillman, "Speckle in Optical
Coherence Tomography," in Advanced Biophotonics: Tissue Optical Sectioning (V. V. Tuchin, and
R. Wang, eds), Taylor & Francis: 211-277 (2013).
Other publications
R. A. McLaughlin, B. C. Quirk, D. Lorenser, X. Yang, B. Y. Yeo, A. Curatolo, K. M.
Kennedy, L. Scolaro, R. W. Kirk, and D. D. Sampson, "A microscope in a needle", Optics
and Photonics News, vol. 23, no. 12, p. 40, December 2012. (This article was selected for
the 2012 special issue "Optics in 2012". It also featured in a video highlight of the issue,
https://www.osapublishing.org/opn/abstract.cfm?uri=opn-23-12-40).
B.F. Kennedy, L. Chin, K.M. Kennedy, P. Wijesinghe, A. Curatolo, S. Es’haghian, P.R.T.
Munro, R.A. McLaughlin, and D.D. Sampson, "Optical elastography: a new window into
disease, " Optics & Photonics News, vol. 25, no. 12, December 2014. (This article was
selected for the 2014 special issue "Optics in 2014". http://www.osa-
opn.org/home/articles/volume_25/december_2014/extras/a_new_window_into_disease)
xxvi
List of acronyms
Acronym Definition
1-D One-dimensional
2-D Two-dimensional
3-D Three-dimensional
AFM Atomic force microscopy
ASP Angular spectrum propagation
CPU Central processing unit
CT Computed tomography
CW Continuous wave
DOE Diffractive optical element
DOF Depth of field
DVC Digital volume correlation
EM Electromagnetic
FD-OCT Fourier-domain optical coherence tomography
FDODL Frequency-domain optical delay line
FDTD Finite-difference in the time-domain
FEM Finite-element method
FFT Fast Fourier transform
FOV Field of view
FWHM Full-width at half maximum
xxvii
GPU Graphical processing unit
H&E Haematoxylin and eosin
HMDS Hexamethyldisilazane
ISAM Interferometric synthetic aperture microscopy
LCoS Liquid crystal on Silicon
MEMS Micro-electrical mechanical system
MIP Maximum-intensity projection
MRI Magnetic resonance imaging
NA Numerical aperture
OCE Optical coherence elastography
OCM Optical coherence microscopy
OCT Optical coherence tomography
OFDI Optical frequency domain imaging
ONH Optic nerve head
PDMS Polydimethylsiloxane
PF Phase Fresnel (lens)
PMMA Poly-methyl methacrylate
PSTD Pseudospectral time-domain
PSF Point spread function
PVA Polyvinyl alcohol
PWM Pulse-width modulation
RAM Random access memory
xxviii
ROI Region of interest
RTE Radiative transport equation
RTV Room-temperature vulcanising
SAW Surface acoustic wave
SD-OCT Spectral-domain optical coherence tomography
SF Scattering-free
SLD Superluminescent diode
SLM Spatial light modulator
SBR Signal-to-background ratio
SNR Signal-to-noise ratio
SS-OCT Swept-source optical coherence tomography
SW Shear wave
TD-OCT Time-domain optical coherence tomography
TE Transverse electric
TM Transverse magnetic
UHROCE Ultrahigh-resolution optical coherence elastography
VVG Verhoeff-Van Gieson
WLS Weighted least squares
xf-FDOCM Extended-focus Fourier-domain optical coherence microscopy
1
Chapter 1
Introduction 1
A major challenge in improving patient health and treatment lies in understanding the
origin of disease and how it affects living tissue across different length scales, from the
molecules inside a cell, to the whole organs within the body. Another challenge lies in early
detection of disease, in order to contain its progression and minimise damage.
Medical imaging has enabled great progress in these two areas. No technique, however,
can produce images with resolution sufficient to see cells and reveal the structural and
functional information at depths of more than a few hundred micrometres below the tissue
surface of a living subject, a task that is necessary to provide a viable substitute for
histopathological analysis.
For a pathology laboratory to provide this information, costly tissue excisions or
biopsies are required, followed by extensive and time-consuming preparation for slicing
and staining before performing microscopy investigation. In certain cases, biopsies are not
even feasible, where tissue excision would compromise organ functionality, e.g., in the brain
or in the eye.
Let us consider the eye, perhaps the most demanding example, where microscopic
functional and structural information is vital in the understanding, early diagnosis and
management of a condition like glaucoma. Even though this thesis does not concern
ophthalmology, the following example serves to highlight the need for improved image
quality and performance in high-resolution bio-imaging to be able to clearly identify tissue
types and constituents, and study and unequivocally diagnose disease.
Glaucoma is the second most common cause of blindness worldwide [1] and leads to
vision loss by damaging retinal ganglion cell axons in and around the optic nerve head
(ONH) [2]. The onset of glaucoma is influenced by many factors [3], with ONH
biomechanics being an important driving mechanism of this disorder, as demonstrated by a
large body of research [4, 5]. However, quantifying ONH biomechanics is complex, and so
far investigators have used general analytical [6, 7] or computational models [8, 9],
untailored to individual patients. To understand the influence of ONH biomechanics on
glaucoma, and then achieve clinical utility, in vivo measurements of the geometry and the
2 Chapter 1 Introduction
mechanical properties of all tissues within the ONH, as well as the load (i.e., intraocular
pressure) that acts on them, are required. This information can then be used in patient-
specific biomechanical models of the ONH. Optical coherence tomography (OCT) has the
potential to be a powerful tool for quantification of the in vivo biomechanics of the ONH.
OCT is a three-dimensional structural biomedical imaging technique that provides
higher spatial resolution (1-20 μm) than modalities such as ultrasound, magnetic resonance
imaging (MRI), and X-ray computed tomography (CT) to depths of several millimetres in
tissue [10]. This penetration depth exceeds that of other high-resolution optical imaging
techniques, such as confocal microscopy or non-linear microscopy, as near-infrared light
beams and coherent detection of backscattered light are used.
The newest generation of OCT devices can acquire serial cross-sectional 2-D images
rapidly (50,000+ depth lines/second), yielding near-real time 3-D volumes of tissue with an
axial resolution of approximately 4 μm for commercial devices, improving to 1 μm for
experimental ultrahigh-resolution devices [11]. The structures of the lamina cribrosa and the
adjacent peripapillary sclera [12, 13], both of which are believed to strongly influence ONH
biomechanics [14, 15] are so deep in the eye that light penetration of commercially available
OCT scanners is only sufficient to start visualizing and identifying their upper boundaries.
The principal reason for failing to visualise these deep structures is the influence on the
OCT image quality of blood vessels, in particular those arising from the central retinal
vessel trunk, as they cast signal shadows and alter structural and tissue displacement
measurements. Furthermore, as incident light travels through the ONH, it attenuates with
depth, such that reflected signals from deep structures may be too weak or the image
quality too poor for them to be reliably detected.
Image quality is defined by descriptors, such as resolution, depth of field (DOF),
signal-to-noise ratio (SNR), sensitivity, contrast, penetration depth and speckle contrast.
These parameters can be severely affected by the alteration of the optical beam (phase and
amplitude) as it propagates through turbid media, resulting in a degradation of the image
quality. These alterations are mainly caused by sample refractive index inhomogeneities
present in closely packed tissue constituents or at tissue boundaries. Degradation can take
the form of tissue-induced aberrations, caused by large-scale refractive index fluctuations,
or diffused haze on the image brought upon by either small-angle multiply-scattered light
wavefronts or wide-angle multiply-scattered light wavefronts that carry little or no
structural information about the sample under investigation.
The intrinsic contrast provided by scattering from the tissue reveals many
morphological features, but is, on occasion, insufficient. For example, it does not readily
reveal microvasculature, and it can be difficult to distinguish tissue types. Mechanical
1.1 Research objectives 3
properties are important to measure in their own right, but they also represent an
alternative form of contrast to optical properties, which provides new opportunities in
imaging tissues. Elastography is a medical imaging technique based on measuring the
spatially resolved response of tissue to mechanical loading and can provide a map of
mechanical properties. Better resolution than manual palpation can be achieved with
elastography based on magnetic resonance imaging or ultrasound imaging [16]. These types
of elastography are emerging as a clinical tool in the diagnosis liver fibrosis and breast
cancer [17]. With resolution of tens of micrometres, optical coherence elastography (OCE)
[18], a form of elastography based on OCT, shows promise in visualizing mechanical
contrast in tissue on a scale intermediate between that of cells and organs.
This resolution is yet too coarse to probe changes to the mechanical properties of tissue
on the cellular scale (< 25 µm), as is ideally required to study the onset and development of
disease [19], preventing OCE from realising its potential as a promising and unique
instrument in the field of cell mechanics. The field of cell mechanics, in part, focuses on
pathogenesis studies at the cellular scale and on the characterisation of the mechanical
signatures of healthy and diseased cellular tissue constituents [20]. Ideally, such studies
would include the ability to provide in situ images at cellular resolution in live tissues in their
native environment.
However, despite the efforts of the scientific community to improve the system
resolution and increase the signal depth penetration, there are severe limitations when using
OCT to image deep tissue structures and when using OCE to resolve tissue mechanical
interaction on the cellular scale. Such limitations lead to artefacts, which may result in
clinical misinterpretation and morphometric errors, or low sensitivity (high rate of false
negative detection of tissue that is actually diseased) and low specificity (high rate of false
positive detection of tissue that is actually healthy). Strongly scattering and attenuating
structures (e.g., pigment and blood, generally highly forward scattering) adversely affect
biomedical OCT applications, by reducing SNR, contrast and resolution. OCT phase
decorrelation noise, limited depth of field, low scan rates or coarse displacement estimation
methods have so far prevented the use of OCE in an ultrahigh-resolution regime.
Since, for the study and diagnosis of diseases, the ultimate goal is to measure
morphometric and biomechanical parameters, it is crucial to obtain high-quality images of
the structures of interest at depth below the surface. The challenges previously mentioned
can be overcome using particular approaches. Amongst these, one is using software post-
processing techniques and algorithms to improve the quality of OCT images [21]. A
potentially more effective one is using hardware techniques, and the use of optical imaging
simulations to guide the hardware development. This is because, in general, a model and
4 Chapter 1 Introduction
the results of a simulation inform the experiments and help advance understanding of the
technique, which remains relatively weak in OCT, and is embryonic in OCE.
The aim of this thesis is to address these issues with hardware techniques, in order to
maximise the potential of OCT and OCE imaging at high resolution in turbid tissues. We
shall not focus on a particular application, as we want to keep a general approach, suited to
most imaging applications of OCT or OCE in turbid biomedical tissues. Therefore, we
want to concentrate on the technological push within these boundaries, with the goal of
significantly improving the microscopic detection of diseased tissue.
1.1 Research objectives
The focus of this PhD thesis is to understand, characterise and improve image quality in
OCT and OCE, performed at millimetre depths in biological tissue, by means of both
experimental and theoretical approaches: optical beam shaping and optical simulations.
We plan to do so, by addressing fundamental limitations of coherent light-tissue
interaction and technical constraints, affecting OCT image quality, with the objective of
opening up new avenues for applications. The research required to achieve this goal can be
broken down into three parts:
1. Providing the tools needed to improve and quantify OCT and OCE image
quality in turbid tissue, i.e., the tools to alter it (beam shaping), measure it (tissue
phantoms), and analyse it (simulations);
2. Analysing OCT image quality in turbid tissue, altered using Bessel and Gaussian
beams, and measuring the relative improvements;
3. Improving OCE image resolution without compromising other image quality
descriptors (e.g., depth of field, strain sensitivity), in order to perform OCE at
the cellular level, with applications in cell mechanics in situ.
1.2 Structure of the thesis
The content of each chapter is briefly summarised below. Where journal papers are
included in the chapters, they are reproduced as published, as noted in this introduction.
Where chapter sections are only partially based on journal papers or other sources, proper
referencing is also listed in the following breakdown.
1.2 Structure of the thesis 5
Chapter 2 - Background of OCT and OCE
This chapter provides a brief overview of OCT technology, concepts and characteristics
that we exploit in our research effort to improve image quality and diagnostic ability in
biological tissue. We review OCT, its working principles and image quality descriptors,
such as resolution, DOF, SNR, sensitivity, contrast, speckle and present an introduction to
OCE, an OCT-based modality for imaging tissue elasticity [18, 22, 23]. Section 2.2 is based
on the introduction of [24]. Section 2.2.2 is based on Section 2 of [25]. Section 2.3.3 is
based on Section 6.1 of [26].
Chapter 3 - Energy-efficient Bessel beams: a tool to alter image quality
This chapter explores the characteristics of differently shaped optical beams used to
interrogate biological tissue and form an image. We present different ways of shaping an
optical beam, with refractive and diffractive optical elements, and with the use of
reconfigurable beam shapers, such as a phase-only liquid crystal-on-silicon spatial light
modulator (SLM). With appropriate characterisation, programming and placement in the
sample arm optical path, we obtained phase-stable, achromatic and diffraction-efficient
Bessel-like beams, and characterised their use in OCT, including their energy efficiency.
Section 3.2 reports the journal paper [27] as published. In this section, we show that,
for quasi non-diffracting Bessel beams, the Fresnel number is the key parameter
determining the trade-off between DOF extension and OCT sensitivity loss when
compared to a Gaussian beam of equal resolution. Section 3.3 is based on the experiment
section of the supplementary information of [28]. We conclude this chapter by discussing
the implication of the so-called self-reconstructing property of the Bessel beam in turbid
tissue imaging.
Chapter 4 - Structured phantoms: a tool to mimic turbid tissue and compare image
quality
This chapter introduces tissue-mimicking targets, called phantoms, that reproduce the
optical, mechanical and structural properties of biological tissue, and can be used to
characterise OCT image quality. Section 4.1 is based on the introduction of [29]. Section
4.2 is based on Section 6.5 of [26], and it reviews the basics of light-tissue interaction, the
different categories of OCT signal contributions from different scattering regimes and their
implications for image quality. We then review Mie theory, describing the main tissue
optical properties, scattering coefficient 𝜇𝜇𝑠𝑠, and scattering anisotropy, 𝑔𝑔. Those properties
6 Chapter 1 Introduction
influence the OCT signal, with respect to both its attenuation with depth and its contrast.
This section is partly based on Sections 2.1.2 and 2.1.3 of [29]. Section 4.3 describes and
validates the fabrication of phantoms with controlled attenuation coefficients 𝜇𝜇𝑡𝑡, and
scattering anisotropy, 𝑔𝑔 and is partly based on Section 2.2 of [29] and the beam and sample
configuration section of the supplementary information of [28]. Section 4.4, based on
Section 3 of [29], discusses and validates fabrication of phantoms with controlled
mechanical properties, especially stiffness. Section 4.5 reports the journal paper [30] as
published, describing novel phantoms containing three-dimensional structure, suitable for
mimicking the complexity of tissue structures on a scale intermediate between the OCT
system resolution and the field of view, and their potential application in the assessment of
speckle. In Section 4.6, based on the beam and sample configuration and validation
sections of the supplementary information of [28], we look more specifically at the use of
phantoms to test OCT image quality descriptors, including, resolution and contrast, with
both a nanoparticle-embedded phantom and an advanced version of a 3-D-structured
phantom. In Section 4.7, based on Section 2.2. of [24], Section 4.1 of [25] and partly [31],
we describe the fabrication and use of structured phantoms for OCE. We conclude the
chapter with a discussion of the use and limitations of silicone phantoms as tissue-
mimicking objects and image test targets.
Chapter 5 - Simulation of beam propagation and image formation in turbid
samples: a tool to theoretically quantify image quality
This chapter presents a suite of OCT image simulation and optical beam propagation tools
for the study and understanding of speckle phenomena in OCT, phase-sensitive OCT
measurements, and assessment of the quality of images of scattering turbid media.
We proceed through increasing complexity and realism of the model on which the
simulations are based, and increasing dimensionality from one to three dimensions. Section
5.2 deals with image formation in the single-scattering regime. In Section 5.2.1, based on
Section 6.2 of [26], we start by simulating the OCT signal from a multitude of scatterers in
one dimension and in two dimensions as the result of local sums of random phasors, and
in Section 5.2.2, based on Section 3.1 of [25], we explore a more advanced linear systems
model providing realistic OCT amplitude and phase images from numerical scattering
phantoms. Section 5.3 presents our journal paper [32], as published, on a two-dimensional
full wave model of image formation in OCT applicable to general samples. Section 5.4,
based on the supplementary information of [28], extends the simulation of the beam
propagation into the sample to three dimensions.
1.2 Structure of the thesis 7
Chapter 6 - Quantifying OCT image quality in turbid samples using Gaussian and
Bessel beams
In this chapter, we answer the question of how energy-efficient Bessel beams perform in
turbid tissue OCT imaging, and whether they hold an advantage in terms of image quality
compared to imaging with Gaussian beams.
Light interaction with turbid tissue affects the image quality indicators. In fact, the
presence and relative contribution of the different categories of image-carrying and
image-degrading light in the detected OCT signal determines the precision and ability to
localise and discriminate the position and intensity of the backscattering events generated
by the tissue refractive index distribution.
In our effort to benchmark image quality and to provide strategies for improvement,
we present our work done in quantifying the influence of Bessel beams on image quality in
OCT. This analysis will bring together all the tools developed in the previous chapters.
Firstly, the beam shaping platform used to produce Gaussian and Bessel beams of equal
transverse resolution. Secondly, the overlayers and structured phantoms to test different
realistic scattering conditions and to quantify the image quality. Finally, the 3-D beam
propagation simulation to verify the different image-carrying and image-degrading light
contributions to the OCT signal. These tools will help us answer the question of which
beam type attains better OCT image quality in turbid tissue under general tissue-like
scattering conditions. Section 6.2 reports our recent journal paper [28], as published.
Chapter 7 - Improving OCE image quality: strain precision and resolution
This chapter focuses on the characterisation and improvement of optical coherence
elastography image quality. We present the results of the optical simulations introduced in
Chapter 5, and incorporate those in a multiphysics simulation, combining optical and
mechanical models, to highlight the influence of acquisition parameters, such as the loading
conditions (compression amplitude), on the elastogram image quality. We do so with
specific reference to elastogram precision, i.e., the repeatability of the tissue displacement
and strain measurement, as determined by indicators such as displacement and strain
sensitivity and strain SNR. Section 7.2 includes most of the journal paper [25] on this
subject.
With the goal of expanding the capabilities of OCE to serve the field of cellular
biomechanics in situ, we then concentrate on improving the elastogram resolution, without
compromising its precision. We do so, by designing an ultrahigh-resolution optical
8 Chapter 1 Introduction
coherence microscopy system with an advanced beam shaping setup, combining Bessel
beam illumination and Gaussian beam detection, and devising an image acquisition strategy
that preserves elastogram precision. This highly novel system and its results are reported in
Section 7.3, where the journal paper [31] is presented as published.
We demonstrate this improvement on both tissue-mimicking phantoms and freshly
excised mouse aorta, revealing the mechanical heterogeneities of vascular smooth muscle
and elastin sheets in the aorta wall in exceptional detail.
Chapter 8 - Conclusions
This chapter summarizes the significance and limitations of the research presented in this
thesis, and makes recommendations for future work. The thesis concludes with a summary
of the key contributions and some final remarks.
9
Chapter 2
Background of OCT and OCE 2
2.1 Optical coherence tomography
This chapter provides a brief overview of the imaging technology, concepts and
characteristics required to understand the context and our effort to improve image quality
and diagnostic ability in biological tissue. We will review OCT, its working principles and
imaging properties, such as resolution, depth of field, signal-to-noise ratio, sensitivity,
contrast, speckle and presents an introduction to optical coherence elastography (OCE), an
OCT-based modality for imaging tissue elasticity [18, 22, 23].
OCT is an optical imaging technique acquiring three-dimensional (3-D), high-
resolution, near real-time images of samples and their sub-surface structure, suitable for
biomedical diagnostics in vivo [33]. It is non-ionizing and non-invasive as it uses infrared
light without requiring direct contact between probe and tissue.
OCT is analogous to ultrasound imaging in its working principle, as it gates the
detected backscattered radiation from the sample and determines the associated distance
from the measured echo-time delay, but uses light waves instead of sound waves. Direct
time-of-flight measurement of the reflected waves is infeasible, since the speed of light is
roughly six orders of magnitude faster than the speed of sound. Instead, low-coherence
interferometry [34, 35], explained in detail in Section 2.1.1, is used to make depth-resolved
measurements of the sample’s reflectance. Light from a spectrally broadband source is split
into two optical paths (‘arms’), and the interference of light reflected or backscattered from
the sample with light reflected from a reference mirror is detected. The use of
low-coherence interferometry for in vivo measurements of tissue has been demonstrated
since at least the mid 1970s [36]. It wasn’t until 1991 that Huang et al. presented the first
demonstration of OCT, producing images by raster scanning the sample beam of a low
coherence interferometer over ex vivo tissue samples [37].
In OCT, a 3-D image is usually formed by scanning in the following order: a one-
dimensional (1-D) axial scan in depth, 𝑧𝑧, is performed by low-coherence interferometry.
10 Chapter 2 Background of OCT and OCE
Acquiring multiple 1-D scans at different transverse locations along 𝑥𝑥 leads to a two-
dimensional (2-D), cross-sectional scan. A 3-D volume image is then formed by scanning
laterally in 𝑦𝑦, and acquiring multiple 2-D scans at different transverse locations. Using the
same terminology as used in ultrasound imaging, the 1-D depth scan is called an A-scan,
the 2-D scan is called a B-scan, and the 3-D scan is called a C-scan. Once acquired, the data
volume can then be digitally sliced in different orientations, similarly to multiplanar
reformatting in CT. Slices in the 𝑥𝑥𝑦𝑦-plane, thus at some depth 𝑧𝑧, are referred to as the
en-face images. Figure 2-1 shows a schematic of this geometry and terminology.
Figure 2-1. Schematic of terminology used when referring to OCT scan orientation. (a) A-scan: a 1-D axial scan formed from the irradiance of light vs. depth, z. (b) B-scan: a 2-D cross-sectional scan formed by scanning laterally (x) and acquiring multiple A-scans. (c) C-scan: a 3-D volume formed by scanning laterally (y) in the orthogonal direction to (b), and acquiring multiple B-scans. Adapted from [11].The resolutions (in the axial and transverse
directions) of a typical OCT system (at focus and in low-scattering media) are around 5–20
μm [11], but systems with ultrahigh resolutions of 1–3 μm have been demonstrated [38].
Unlike in most other microscopy techniques, the mechanisms determining the axial and
transverse resolution in OCT are decoupled, as only the transverse resolution is determined
by the sample optics, while the axial resolution is determined by the coherence property of
the light source. Section 2.3.1 deals in detail with the factors influencing OCT resolution.
Light extinction from tissue is caused by absorption and scattering processes. The
wavelengths used for OCT are usually chosen to lie within the so-called diagnostic window
(650–1,350 nm) to minimise the absorption of light by tissue constituents [39, 40]. Figure
2-2 shows a diagram of the spectral molar extinction by selected tissue chromophores.
2.1 Optical coherence tomography 11
Figure 2-2. Spectral absorption for a range of tissue chromophores in the diagnostic window in the near-infrared. For wavelengths in the range 650–1350 nm, the extinction of light due to absorption and scattering is relatively low. Based on data from [41-44].
The optical absorption of any given chromophore in tissue is determined by the molar
extinction coefficient multiplied by the concentration of that chromophore. Tissue
absorption is dominated by the spectral response of water since water is the most
prominent constituent of most soft tissues. Thus, water limits light penetration in tissue on
the long wavelength side, and melanin and blood, where present, on the short wavelength
side. Within the diagnostic window, light scattering is two or more orders of magnitude
stronger than absorption, making it very suitable for OCT, which relies on the elastic
backscattering of light to form an image. The most commonly used wavelengths for OCT
are centred around 800 nm, which provides higher resolution (for a given bandwidth, see
Section 2.3.1), and 1,300 nm, which provides greater depth penetration in tissue. The
typical depth penetration of OCT is in the range 0.5–1.5 mm, determined primarily by the
interplay between scattering, system sensitivity and numerical aperture.
In terms of resolution and penetration depth, OCT occupies a niche between
laboratory imaging techniques, such as confocal microscopy, and medical imaging
techniques, such as ultrasound and magnetic resonance imaging, as illustrated in Figure 2-3.
12 Chapter 2 Background of OCT and OCE
Figure 2-3. Comparison of OCT resolution and imaging depths to other biomedical imaging methods. The “pendulum” length represents imaging depth, and the “sphere” size represent resolution. Reproduced from [45].
OCT is clinically used primarily in ophthalmology [46, 47], especially for imaging the
retina, but increasingly also the anterior segments of the eye, such as the cornea. OCT is
also being used clinically, in order of decreasing importance, for cardiology [48, 49],
dermatology [50], gastroenterology [51], dentistry [52], and pulmonology [53, 54], as shown
in Figure 2-4. Examples of the three most published research application areas are
schematically shown in Figure 2-5. Figure 2-6 shows movies of representative OCT images
from those clinical applications as well as laboratory applications.
The advances into fibre-optics technology brought by the telecommunications industry
had fostered the growth of OCT imaging and helped overcome its short penetration depth
by delivering light through flexible probes, allowing for in vivo and in situ imaging of internal
tissues.
Fibre-optics and miniaturised optics are at the core of compact endoscope [55],
catheter [56] and needle [57] probes. In addition, OCT has been used as the basis for
several derived modalities, including Doppler OCT for measuring fluid velocities,
particularly of blood flow in vasculature [58-60], and spectroscopic OCT for measuring
depth-resolved, wavelength-dependent attenuation [61].
2.1 Optical coherence tomography 13
Figure 2-4. Bar chart of a snapshot of results from a Scopus search of selected OCT biological applications from 2005 to 2015, inclusive. Searches were conducted on the publication title and only included English results. Search terms used were as follows: “optical coherence tomography” AND terms relevant for each heading, e.g. "optical coherence tomography" AND (breast OR mamm*), where the use of a * indicates a search for all words containing the truncated term. (http://www.scopus.com).
Figure 2-5. Main applications of OCT. (a) Ophthalmology. (b) Cardiology. (c) Dermatology. Reproduced from [62-72].
13152135365682103110153
5612736
0 500 1000 1500 2000 2500 3000
GynaecologyLaryngology
MuscleBreast
UrologyGastroenterology (inc. pancreas and biliary tract)
Pulmonary medicineDentistry
Neurology (exc. ophthalmology)Dermatology
CardiologyOphthalmology
# of publications between 2005 and 2015
# of publications
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