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Article in Proceedings of SPIE - The International Society for Optical Engineering · February 2014
DOI: 10.1117/12.2039576
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ABSTRACT
Molecular profiling of tissue using near-infrared (NIR) Raman spectroscopy has shown great promise for in vivo
detection and prognostication of cancer. The Raman spectra measured from the tissue generally contain fundamental
information about the absolute biomolecular concentrations in tissue and its changes associated with disease
transformation. However, producing analogues tissue Raman spectra present a great technical challenge. In this
preliminary study, we propose a method to ensure the reproducible tissue Raman measurements and validated with the in
vivo Raman spectra (n=150) of inner lip acquired using different laser powers (i.e., 30 and 60 mW). A rapid Raman
spectroscopy system coupled with a ball-lens fiber-optic Raman probe was utilized for tissue Raman measurements. The
investigational results showed that the variations between the spectra measured with different laser powers are almost
negligible, facilitating the quantitative analysis of tissue Raman measurements in vivo.
__________________________________________________________________________________
*Corresponding to: Dr Zhiwei Huang, E-mail: biehzw@nus.edu.sg, Tel: +65 6516 8856, Fax: +65 6872 3069
Optical spectroscopy methods that can probe changes in tissue morphology, intrinsic fluorophores and biomolecules are
promising candidates suited for biomedical diagnosis. Among various optical spectroscopic methods, fiber-optic Raman
spectroscopy is a unique analytical technique, and has recently opened a new pathway for fighting against cancer based
on early detection by providing quantitative and quality information of tissue composition [1-3]. The advancements in
modern Raman instrumentation, recent breakthroughs in the development of miniaturized fiber-optic Raman probe
designs, and robust multivariate data processing methods have opened a new frontier of real-time Raman spectroscopy
for early cancer screening in vivo in a number of organs (e.g., bladder, nasopharynx and larynx, cervix, lung, esophagus,
and stomach, etc) [2, 4-9]. The biomedical tissue samples are often inhomogeneous and the acquired tissue Raman
signals are extremely complex, containing absolute biomolecular concentration information in the tissue and its changes
associated with disease transformation. Thus the quantitative Raman spectroscopy has the promising potential to greatly
improve tissue diagnosis. However, producing analogues Raman spectra from inhomogeneous tissue present a great
technical challenge to the biomedical community that requires controlling and monitoring of the most common sources
of variances in fiber-optic quantitative analysis, including laser excitation power, fiber coupling efficiency and
measurement reproducibility, etc. [10]. Normalization to the peak intensity or area under the curve is the most
widespread method to detect the disease based on the spectral shape information, while disregarding the quantitative
(intensity) information [11]. Till date, very limited studies have been reported on quantitative tissue Raman spectroscopy
using fluorocarbon fibers, a fluorinated ethylene propylene copolymer cap on the excitation fiber of Raman probes, and
fiber-optic Raman probes with embedded diamond in the fiber-tip or distal lens [10, 12-14]. In this study, without
inserting additional reference materials or impurities into the fiber-optic Raman probe, we utilize sapphire Raman signals
generated from the distal ball-lens of confocal fiber-optic Raman probe as an internal reference for quantitative fiber-
optic tissue Raman measurements.
2.2 Patients
A total of 15 normal healthy volunteers between 18 to 35 years of age were recruited for in vivo tissue Raman
measurements in the oral cavity. Prior to research investigation, all patients signed an informed consent form permitting
the collection of in vivo spectroscopic measurements from the oral cavity. Before taking the spectroscopic measurement,
all volunteers underwent extensive mouthwash to reduce confounding factors (e.g., food debris, microbial coatings etc.).
As a result, a total of 150 in vivo tissue Raman spectra were measured from the inner lip of volunteers, in which 80
spectra were acquired using laser power of 60 mW and 70 spectra were measured using the laser power of 30 mW.
3. Results
Figure 1 shows the background spectrum of a ball-lens fiber-optic Raman probe under the 785 nm laser excitation with a
laser power of ~30 mW. The Raman peaks of sapphire (Al2O3) distal ball-lens can be found at 417 and 646 cm-1 (phonon
mode with A1g symmetry), and 380 and 751 cm-1 (Eg phonon mode). The AF background and the Raman peaks arising
from fused silica fibers likely due to the metal ion impurities in the silica fibers were also noticed around 490 and
606 cm−1 (Figure 1) [10].
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Fig.1. Background Raman spectrum of a ball-lens fiber-optic Raman probe when excited by a 785 nm laser
with an excitation power of ~30 mW.
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4. CONCLUSION
This study demonstrates that the internal reference Raman signals arising from the ball-lens of fiber-optic Raman probes
can be utilized for realizing quantitative analysis of tissue Raman measurements in vivo.
ACKNOWLEDGEMENTS
This work was supported by the National Medical Research Council and the Biomedical Research Council, Singapore.
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