This PDF file contains the front matter associated with SPIE Proceedings Volume 7170, including the Title Page, Copyright information, Table of Contents, Introduction (if any), and the Conference Committee listing.

We present a new type of protein microarray called the Land-contrast (LC) BioCD in which imaging contrast is induced by a patterned substrate rather than by patterned protein. This is realized by etching spot patterns in a silicon wafer. On the spot region the silicon dioxide thickness is 140 nm and on the land it is 77 nm. The spot and the land have equal reflectance but opposite interferometric quadrature responses for protein layer. Protein is evenly immobilized on the entire chip and detected by reflectometry. Therefore there is no need for protein printing, nor spectrometers, nor high angles nor polarization control to image the surface-bound protein. The LC BioCD can facilitate research on protein microarrays.

It has been shown that frequency domain optical coherence tomography (FD-OCT) systems achieve higher sensitivities compared to time domain OCT systems. However, the obscure object structure due to the mirror image generated by the Fourier transform is one of the main problems in FD-OCT. We designed and developed a novel full range FD-OCT system that we refer to as the dual-reference full-range FD-OCT (DR-FDOCT) that enables doubling the imaging depth by removing mirror images generated from the Fourier transform of the detected real-valued spectra. The DR-FDOCT system enables full-range imaging without any loss of speed and is less sensitive to phase errors generated by involuntary movements of the subject than the other established full-range OCT systems. The reason is because it uses two signals with a phase difference of π/2 obtained simultaneously from two reference arms to remove mirror images.

Non-contact reflection photoplethysmography (NRPPG) is being developed to trace pulse features for comparison with contact photoplethysmography (CPPG). Simultaneous recordings of CPPG and NRPPG signals from 22 healthy subjects were studied. The power spectrum of PPG signals were analysed and compared between NRPPG and CPPG. The recurrence plot (RP) was used as a graphical tool to visualize the time dependent behaviour of the dynamics of the pulse signals. The agreement between NRPPG and CPPG for physiological monitoring, i.e. HRV parameters, was determined by means of the Bland-Altman plot and Pearson's correlation coefficient. The results indicated that NRPPG could be used for the assessment of cardio-physiological signals.

The work described in this presentation relates to a novel approach to Electron Paramagnetic Resonance imaging in vivo. The method employs a continuous wave approach to spectral detection without the conventional low frequency modulation and phase sensitive detection. A combination of direct detection and rapid field scan in the simultaneous presence of rotating gradients enables imaging with high temporal resolution. Since CW detection is not limited to free radical spin probes with narrow spectral lines, unlike the time-domain case, this novel approach uniquely accomplishes ultra fast functional imaging and is applicable to common redox-sensitive spin probes without line-width restriction.

Achieving high lateral resolution still remains a challenge for in vivo Optical Coherence Microscopy (OCM) biological imaging. While to address this challenge, the numerical aperture (NA) of the microscope objective in the sample arm of the OCM interferometer may be increased, it introduces trade-offs in terms of loss in the depth of focus over which lateral resolution can still be maintained. As a critical step to offset this problem, we recently presented the optical system design of a dynamic focusing (DF) optical coherence microscope with a built-in liquid lens for re-focusing through the sample depth with no moving parts at in vivo speeds. We present experimental measurements of the modulation transfer function (MTF) acquired from the fabricated research prototype. The measurements were obtained though the edge detection method as a function of the voltage applied and at various positions in the field of view (FOV) within a 2mm cubic sample. Results demonstrate a resolution of 2 µm across the voltage range and the FOV, which validates the expectation by design of a quasi-invariant resolution of less than 3μm over a 2mm×2mm lateral cross-section across the 2mm depth of skin-equivalent tissue. Images of a tadpole sample acquired with the probe at different focal depths are also shown to demonstrate gain in resolution with focusing through different depth zones.

Reflectance confocal microscopy provides real time, cellular resolution images of in-vivo and ex-vivo tissues and been cleared by the FDA and international regulatory agencies for medical applications. Clinical applications of reflectance confocal microscopy are being tested in single- and multi-center clinical trials. In this talk I will review the design challenges of sub-surface imaging with confocal microscopy and techniques to compare instrument performance between different sites.

Gastrointestinal endoscopy has made great progress during last decade. Diagnostic accuracy can be enhanced by better training, improved dye-contrast techniques method, and the development of new image processing technologies. However, diagnosis using conventional endoscopy with white-light optical imaging is essentially limited by being based on morphological changes and/or visual attribution: hue, saturation and intensity, interpretation of which depends on the endoscopist's eye and brain. In microlesions in the gastrointestinal tract, we still rely ultimately on the histopathological diagnosis from biopsy specimens. Autofluorescence imaging system has been applied for lesions which have been difficult to morphologically recognize or are indistinct with conventional endoscope, and this approach has potential application for the diagnosis of dysplastic lesions and early cancers in the gastrointestinal tract, supplementing the information from white light endoscopy. This system has an advantage that it needs no administration of a photosensitive agent, making it suitable as a screening method for the early detection of neoplastic tissues. Narrow band imaging (NBI) is a novel endoscopic technique which can distinguish neoplastic and non-neoplastic lesions without chromoendoscopy. Magnifying endoscopy in combination with NBI has an obvious advantage, namely analysis of the epithelial pit pattern and the vascular network. This new technique allows a detailed visualization in early neoplastic lesions of esophagus, stomach and colon. However, problems remain; how to combine these technologies in an optimum diagnostic strategy, how to apply them into the algorithm for therapeutic decision-making, and how to standardize several classifications surrounding them. 'Molecular imaging' is a concept representing the most novel imaging methods in medicine, although the definition of the word is still controversial. In the field of gastrointestinal endoscopy, the future of endoscopic diagnosis is likely to be impacted by a combination of biomarkers and technology, and 'endoscopic molecular imaging' should be defined as "visualization of molecular characteristics with endoscopy". These innovations will allow us not only to locate a tumor or dysplastic lesion but also to visualize its molecular characteristics (e.g., DNA mutations and polymorphisms, gene and/or protein expression), and the activity of specific molecules and biological processes that affect tumor behavior and/or its response to therapy. In the near future, these methods should be promising technologies that will play a central role in gastrointestinal oncology.

Two different, already characterized, hyperspectral imaging systems created for visualizing the spatial distribution of tissue oxygenation non-invasively for in vivo clinical use are described. Individual components of both liquid crystal tunable filter (LCTF) and digital light processing (DLP) systems were characterized, calibrated, and found to be well within manufacturer specifications. Coupling LCTF with charge coupled device (CCD) technology and acquiring images at multiple, contiguous wavelengths and at narrow bandwidths are formatted into a hyperspectral data cube consisting of one spectral and two spatial dimensions. DLP® technology has the novel ability to conform light to any desired spectral illumination scheme. Subsequently the collected multispectral data are processed into chemically relevant images that are color encoded at each pixel detector for the relative percentage of oxyhemoglobin. Using spectral illumination methods unique to the DLP hyperspectral imager results in producing chemically relevant images at near video rate; 4 frames per second. As an example, both systems are used to collect spectral data from a 27.22 kg porcine kidney whose renal artery has been occluded for 60 minutes. Both systems return nearly identical spectra collected from the surface of the kidney, with a root mean square deviation between the two spectra of 0.02.

Multi-infusion setups for medication administration in Intensive Care Units seem uncontrolled due to flow and pressure differences between syringe pumps. To investigate the dynamics and interaction of multi-infusion, a dedicated set-up was developed to measure the concentrations of fluids dynamically in multiple lines using absorption spectral-photometry. For feasibility testing and calibration, various dyes and concentrations were investigated to find the optimal settings. The developed method was validated and showed satisfactory results for determining mixtures of up to three different dyes in different ratios, with average recoveries of 105.0% (±11.01) for two dyes and 99.6% (±6.26) for three dyes. The method was applied in initial simulation experiments for measuring effects of manipulations in a multi-infusion set-up simulating a clinical situation. Results showed evidence for mutual influencing of separate infusion lines. The method developed for measuring the fluid dynamics of multi-infusion will contribute to a better insight and controlled administration of medications.

Everyone would like to calibrate their fluorometers with a stable transferable fluorescent object. Existing stable fluorophores generally do not match the geometrical, spectral or anisotropic properties of samples used in in-vitro diagnostics. This paper will discuss the ideal features of fluorescence calibrators and why no practical calibrators exist. Finally, we will suggest partial solutions to the problem.

As sophisticated optical imaging technologies move into clinical applications, manufacturers have to work according to a consistent quality management. We demonstrate the application of basic quality principles to camera-based biomedical optics for a variety of examples including molecular diagnostics, dental imaging, ophtalmology and digital radiography. Novel concepts in fluorescence detection and structured illumination will also be highlighted.

In order to elucidate light propagation mechanisms involved in optical spectroscopy devices, the optical properties of layered mucosal tissues at ultraviolet and visible wavelengths are needed. Previous approaches to measuring this data have typically been based on spatially-resolved reflectance. However, these approaches have limitations, some of which are not well understood. Therefore, the objectives of this study were (1) to elucidate the relationship between spatially-resolved reflectance distributions and optical properties in two-layer tissue models and (2) introduce and assess an unconstrained approach to optical property measurement. The first part of this study involved calculating reflectance from two-layer tissue for a wide variety of optical property combinations (πa = 1-22.5, πs' = 5-42.5 cm-1) using a Monte Carlo scaling technique. In the second part, a neural network inverse model trained with the aforementioned results was evaluated using simulated reflectance data. This relationship between optical properties and reflectance provides fundamental insights into the strengths, weaknesses and potential limitations of strategies for optical property measurement based on spatially-resolved reflectance. The neural network approach estimated optical property values with a degree of accuracy that depended on the probe geometry (5-, 6-, 10- and 11-fiber probes were simulated). The average error in determination of πa ranged from 15 to 51% and average error for πs' ranged from 8 to 32%. While computationally expensive to develop, neural network models calibrated with simulation data may prove to be a highly effective approach for rapid, unconstrained estimation of the optical properties of two-layer tissues.

Three sensor designs for full field laser doppler blood flow imaging will be demonstrated(i) 16x1 pixel linear array with mixed analogue and digital signal processing (ii) 4x4 pixel array with all analogue processing (iii) hybrid system which combines 32x32 pixel array with part on-chip processing linked to a field programmable gate array. Results are demonstrated using modulated light, tissue phantoms and blood flow in tissue. The efficient use of silicon when implementing the signal processing on-chip is discussed.

Near-Infrared (NIR) picosecond pulsed light shined in biological tissues (e.g. brain, breast, muscle) offers the opportunity for non-invasive quantitative spectroscopy and imaging. Tissue optical properties determine high attenuation levels of optical signals and nanosecond scale dynamics. Therefore high-performance set-ups are needed. We aimed at developing a winning photodetector-electronics pairing for a broad field of multiple-wavelengths faint-signal optical investigations, like brain functional imaging, optical mammography, in-vivo spectroscopy, drugs characterization, molecular imaging. We present an electronic instrumentation based on silicon Single-Photon Avalanche Diode (SPAD) and fast-gating frontend electronics, in a Time-Correlated Single-Photon Counting (TCSPC) set-up. Detection efficiency is very high (50% at 550 nm and 15% at 800 nm), allowing acquisition of very faint optical signals on a wide spectral range. Furthermore, the fast-gating circuitry enables the detector very quickly (500 ps) and for user-selectable (200 ps - 510 ns) durations, thus allowing the rejection of very intense optical signals (e.g. scattered light from more superficial layers of the tissue under investigation) preceding useful faint signals (e.g. scattered light from sub-cellular components or coming from "deep" tissue layers), which would be otherwise overwhelmed and made undetectable. We attain photon-counting dynamic ranges up to 107 with photon-timing resolutions of 95 ps.

Time-correlated single photon counting (TCSPC) is popular in time resolved techniques due to its prominent performance such as ultra-high time resolution and ultra-high sensitivity. However, this technique is limited by low counting rate and high system cost. In this paper, we report a new time-resolved optical measurement method which aims to achieve faster data acquisition without losing the key benefits of TCSPC. The new method is based on the spread spectrum time-resolved optical measurement method combined with single photon counting. A pseudo-random bit sequence is used to modulate a continuous wave laser diode, while the pulse sequence in response to the modulated excitation is recorded by a single photon detector. The impulse response is then retrieved by periodic cross-correlation. Both simulation and experimental work have been conducted to validate our approach. Experimental results with our prototype have shown a time-resolution better than 200 picoseconds. Besides the faster data acquisition and high timeresolution, the new method also affords other benefits such as portability and low cost.

Light is a powerful tool for the life sciences. High intensity, low cost light engines are therefore essential to the design and proliferation of new bioanalytical instruments, medical devices and miniaturized analyzers. Lumencor has developed an inexpensive lighting solution, uniquely well suited to the production of safe, effective, commercial life science devices. Lumencor’s proprietary technology provides powerful, pure, stable, inexpensive light across the UV-Vis-IR. Light engines are designed to directly replace the entire configuration of light management components with a single, simple unit. Multicolor prototypes will be discussed and their performance capabilities disclosed.

We report a new close-loop feedback control method to keep a Mach-Zehnder electro-optic modulator (MZ-EOM) biased at the quadrature point and simultaneously correct the bias drift caused by the temperature changes as well as the inherent photorefractive effect. The modulator is a key part of our high speed time-domain diffuse optical tomography system. It modulates the dual-wavelength near-infrared light with the high speed pseudorandom bit sequence (PRBS) signal for the temporal point spread function (TPSF) measurements. Our method applies a periodical low frequency square wave with 50% duty cycle as the pilot tone upon the MZ-EOM together with the PRBS and sweep the bias voltage of the MZ-EOM in a self-adaptive step. A constant fraction of the modulated output power is measured by a photodiode via a tap coupler. After demodulation, the modulation depth versus the bias voltage can be measured from which the peak value corresponding to the quadrature point can be located quickly by curve fitting. Our stabilization technique is simple, fast and cost effective and is effective to correct the bias drift caused by the photorefractive and the change of ambient conditions. The experiment results show the TPSFs measurements can be stabilized to within ±2% in an hour duration, which helps improved the image quality.

Anyone can make a fluorometer. All that is needed is a light source, a detector, some filters and maybe some lenses. For commercial clinical instruments, however, our customers demand stability, reliability, and reproducibility. If research instruments are like race cars, commercial instruments should be like trucks. This demands that our fluorometer designs be carefully toleranced. This paper will discuss causes of variation and drift, and ways to make fluorometers that are stable and reproducible.