Cancer is a major health problem in the world today. Almost all cancers have a
significantly better chance for therapy and recovery if detected at their early stage.
The capability to perform disease diagnosis at an early stage requires high-resolution
imaging that can visualise the physiological and morphological changes at a cellular
level. However, resolving powers of current medical imaging systems are limited
to sub-millimeter sizes. Furthermore, the majority of cancers are associated with
morphological and functional alterations of cells in epithelial tissue, currently assessed
by invasive and time-consuming biopsy. Optical imaging enables visualisations of tissue
microstructures at the level of histology in non-invasive means. Optical imaging is
suitable for detecting neoplastic changes with sub-cellular resolution in vivo without
the need for biopsy.
Nonlinear optical microscopy based on multi-photon absorption and higher harmonic
generation has provided spectacular sights into visualisation of cellular events
within live tissue due to advantages of an inherent sectioning ability, the relatively deep
optical penetration, and the direct visualisation of intrinsic indicators. Two-photon
excited uorescence (TPEF) from intrinsic cell components and second harmonic
from asymmetric supermolecular structures can provide complementary information
regarding functionalities and morphologies in tissue environments, thus enabling
premalignant diagnosis by detecting the very earliest changes in cellular structures.
During the past sixteen years, nonlinear optical microscopy has evolved from a
photonic novelty to a well-established laboratory tool. At present, in vivo imaging and
long-term bedside studies by use of nonlinear optical microscopy have been limited
due to the fact that the lack of the compact nonlinear optical instrument/imaging
technique forces the performance of nonlinear optical microscopy with bulk optics on
the bench top. Rapid developments of fibre-optics components in terms of growing
functionalities and decreasing sizes provide enormous opportunities for innovation in
nonlinear optical microscopy. Fibre-based nonlinear optical endoscopy will be the soul
instrumentation to permit the cellular imaging within hollow tissue tracts or solid
organs that are inaccessible with a conventional optical microscope.
Lots of efforts have been made for development of miniaturised nonlinear optical
microscopy. However, there are major challenges remaining to create a nonlinear
optical endoscope applicable within internal cavities of a body. First, an excitation
laser beam with an ultrashort pulse width should be delivered eciently to a remote
place where ecient collection of faint nonlinear optical signals from biological samples
is required. Second, laser-scanning mechanisms adopted in such a miniaturised
instrumentation should permit size reduction to a millimeter scale and enable fast
scanning rates for monitoring biological processes. Finally, the design of a nonlinear
optical endoscope based on micro-optics must maintain great exibility and compact
size to be incorporated into endoscopes to image internal organs.
Although there are obvious diculties, development of fibre-optic nonlinear optical
microscopy/endoscopy would be indispensible to innovate conventional nonlinear
optical microscopy, and therefore make a significant impact on medical diagnosis. The
work conducted in this thesis demonstrates the new capability of nonlinear optical
endoscopy based on a single-mode fibre (SMF) coupler or a double-clad photonic
crystal fibre (PCF), a microelectromechanical system (MEMS) mirror, and a gradientindex
(GRIN) lens. The feasibility of all-fibre nonlinear optical endoscopy is also
demonstrated by the further integration of a double-clad PCF coupler. The thesis
concentrates on the following key areas in order to exploit and understand the new
imaging modality.
It has been known from the previous studies that an SMF coupler is suitable for twoii
photon excitation by transmitting near infrared illumination and collecting uorescence
at visible wavelength as well. Although second harmonic generation (SHG) wavelength
is farther away from the designed wavelength of the fibre coupler than that of normal
TPEF, it is demonstrated in this thesis that both SHG and TPEF signals can be
collected simultaneously and eciently through an SMF coupler with axial resolution
of 1.8 um and 2.1 um, respectively. The fibre coupler shows a unique feature of linear
polarisation preservation along the birefringent axis over the near infrared and the
visible wavelength regions. Therefore, SHG polarisation anisotropy can be potentially
extracted for probing the orientation of structural proteins in tissue. Furthermore,
this thesis shows the characterisation of nonlinear optical microscopy based on the
separation distance of an SMF coupler and a GRIN lens. Consequently, the collection
of nonlinear signals has been optimised after the investigation of the intrinsic trade-off
between signal level and axial resolution.
These phenomena have been theoretically explored in this thesis through formalisation
and numerical analysis of the three-dimensional (3D) coherent transfer function
for a SHG microscope based on an SMF coupler. It has been discovered that a fibreoptic
SHG microscope exhibits the same spatial frequency passband as that of a fibreoptic
reection-mode non-uorescence microscope. When the numerical aperture of
the fibre is much larger than the convergent angle of the illumination on the fibre
aperture, the performance of fibre-optic SHG microscopy behaves as confocal SHG
microscopy. Furthermore, it has been shown in both analysis and experiments that
axial resolution in fibre-optic SHG microscopy is dependent on the normalised fibre
spot size parameters. For a given illumination wavelength, axial resolution has an
improvement of approximately 7% compared with TPEF microscopy using an SMF
coupler.
Although an SMF enables the delivery of a high quality laser beam and an enhanced
sectioning capability, the low numerical aperture and the finite core size of an SMF
give rise to a restricted sensitivity of a nonlinear optical microscope system. The
key innovation demonstrated in this thesis is a significant signal enhancement of a
nonlinear optical endoscope by use of a double-clad PCF. This thesis has characterised
properties of our custom-designed double-clad PCF in order to construct a 3D nonlinear
optical microscope. It has been shown that both the TPEF and SHG signal levels in
a PCF-based system that has an optical sectioning property for 3D imaging can be
significantly improved by two orders of magnitude in comparison with those in an
SMF-based microscope. Furthermore, in contrast with the system using an SMF,
simultaneous optimisations of axial resolution and signal level can be obtained by
use of double-clad PCFs. More importantly, using a MEMS mirror as the scanning
unit and a GRIN lens to produce a fast scanning focal spot, the concept of nonlinear
optical endoscopy based on a double-clad PCF, a MEMS mirror and a GRIN lens has
been experimentally demonstrated. The ability of the nonlinear optical endoscope to
perform high-resolution 3D imaging in deep tissue has also been shown.
A novel three-port double-clad PCF coupler has been developed in this thesis to
achieve self-alignment and further replace bulk optics for an all-fibre endoscopic system.
The double-clad PCF coupler exhibits the property of splitting the laser power as well
as the separation of a near infrared single-mode beam from a visible multimode beam,
showing advantages for compact nonlinear optical microscopy that cannot be achieved
from an SMF coupler. A compact nonlinear optical microscope based on the doubleclad
PCF coupler has been constructed in conjunction with a GRIN lens, demonstrating
high-resolution 3D TPEF and SHG images with the axial resolution of approximately
10 m. Such a PCF coupler can be useful not only for a fibre-optic nonlinear optical
probe but also for double-clad fibre lasers and amplifiers.
The work presented in this thesis has led to the possibility of a new imaging device
to complement current non-invasive imaging techniques and optical biopsy for cancer
detection if an ultrashort-pulsed fibre laser is integrated and the commercialisation
of the system is achieved. This technology will enable in vivo visualisations of
functional and morphological changes of tissue at the microscopic level rather than
direct observations with a traditional instrument at the macroscopic level. One can
anticipate the progress in bre-optic nonlinear optical imaging that will propel imaging
applications that require both miniaturisation and great functionality.
| Identifer | oai:union.ndltd.org:ADTP/216675 |
| Date | January 2007 |
| Creators | Fu, Ling, n/a |
| Publisher | Swinburne University of Technology. |
| Source Sets | Australiasian Digital Theses Program |
| Language | English |
| Detected Language | English |
| Rights | http://www.swin.edu.au/), Copyright Ling Fu |
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