Biophotonics: a marriage of sciences

Biophotonics denotes a combination of biology and photonics, with photonics being the science and technology of generation, manipulation, and detection of photons, quantum units of light. Photonics is related to electronics in that it is believed that photons will play a similar central role in future information technology as electrons do today. Biophotonics has therefore become the established general term for all techniques that deal with the interaction between biological items and photons. This refers to emission, detection, absorption, reflection, modification, and creation of radiation from biomolecular, cells, tissues, organisms and biomaterials. Areas of application are life science, medicine, agriculture, and environmental science

Use of confocal and multiphoton microscopy for the evaluation of micro-optical components and emitters

We report on the application of confocal and multiphoton microscopic techniques for the evaluation of the latest generation of micro optical components. The optical emitting characteristics of arrays of matrix addressable GaN micrometer-sized light emitting diodes (micro-LEDs) have been measured using a commercial confocal microscope utilising the LEDs' own emission along with reflection confocal microscopy to determine the surface structure. Multiphoton induced luminescence from the 10-20-micron diameter emitters has also been used to examine the structure of the device and we compare this with electrically induced emission. In related work, the optical properties of micro lens arrays (10-100-micron diameter) fabricated in SiC, Sapphire, and Diamond have been determined using transmission confocal microscopy. Such optical microscopy techniques offer a simple, non-destructive method to determine the structure and performance of such novel devices

High-resolution 3D optical microscopy inside the beating zebrafish heart using prospective optical gating

3D fluorescence imaging is a fundamental tool in the study of functional and developmental biology, but effective imaging is particularly difficult in moving structures such as the beating heart. We have developed a non-invasive real-time optical gating system that is able to exploit the periodic nature of the motion to acquire high resolution 3D images of the normally-beating zebrafish heart without any unnecessary exposure of the sample to harmful excitation light. In order for the image stack to be artefact-free, it is essential to use a synchronization source that is invariant as the sample is scanned in 3D. We therefore describe a scheme whereby fluorescence image slices are scanned through the sample while a brightfield camera sharing the same objective lens is maintained at a fixed focus, with correction of sample drift also included. This enables us to maintain, throughout an extended 3D volume, the same standard of synchronization we have previously demonstrated in and near a single 2D plane. Thus we are able image the complete beating zebrafish heart exactly as if the heart had been artificially stopped, but sidestepping this undesirable interference with the heart and instead allowing the heart to beat as normal

Confocal Microscopy using an InGaN violet laser diode at 406nm

We report on the application of a novel all-solid-state violet laser diode source to confocal microscopy. The source has the potential to replace argon ion lasers in a range of fluorescence based imaging systems. Improvements in system performance and image quality through the use of anamorphic prisms to modify the beam profile have been characterised. These modifications have permitted high quality, optically sectioned images to be obtained from laser diodes operating around 406nm. Living mammalian cells stained with a range of biologically significant fluorophores have been imaged. In addition, it has been shown that at this wavelength it is possible to image dyes that normally require excitation with UV argon laser lines

Combining optical trapping in a microfluidic channel with simultaneous micro-Raman spectroscopy and motion detection.

Since their invention by Ashkin optical tweezers have demonstrated their ability and versatility as a non-invasive tool for micromanipulation. One of the most useful additions to the basic optical tweezers system is micro-Raman spectroscopy, which permits highly sensitive analysis of single cells or particles. We report on the development of a dual laser system combining two spatial light modulators to holographically manipulate multiple traps (at 1064nm) whilst undertaking Raman spectroscopy using a 532nm laser. We can thus simultaneously trap multiple particles and record their Raman spectra, without perturbing the trapping system. The dual beam system is built around micro-fluidic channels where crystallisation of calcium carbonate occurs on polymethylmethacrylate (PMMA) beads. The setup is designed to simulate at a microscopic level the reactions that occur on items in a dishwasher, where permanent filming of calcium carbonate on drinking glasses is a problem. Our system allows us to monitor crystal growth on trapped particles in which the Raman spectrum and changes in movement of the bead are recorded. Due to the expected low level of crystallisation on the bead surfaces this allows us to obtain results quickly and with high sensitivity. The long term goal is to study the development of filming on samples in-situ with the microfluidic system acting as a model dishwasher

Differences in optic nerve structure between individuals of predominantly African and European ancestry: Implications for disease detection and pathogenesis

Glaucoma disproportionately affects individual of African ancestry. Additionally, racial differences in the optic nerve head have been well described that may alter the vulnerability to intraocular pressure related injury and, in addition, alter the clinical ability to detect the presence of early optic nerve injury. This paper will review the literature describing racial differences in the optic nerve head between individuals of African and European ancestry with regards to the potential effects of these differences on the ability to detect glaucoma in different racial groups and to potential differences in the pathogenesis of glaucomatous injury

Adaptive Optimisation of Illumination Beam Profiles in Fluorescence Microscopy

Wide-field fluorescence microscope techniques such as single/selective plane illumination microscope (SPIM) are typically configured to image large regions of a sample at once. Here the illumination beam provides uniform excitation of several biological features across the region, `sliced' to a thickness of between 5-10 microns. In this paper we propose a simple alteration to the optical configuration of a SPIM by switching the light-sheet- forming cylindrical lens with a spatial light modulator. This has the potential to adaptively reconfigure the light sheet geometry to improve the optical sectioning of specific biological features, rather than the thicker sectioning of several features at once across a larger observation field-of-view. We present a prototype version of such a system, referred to as an Adaptive-SPIM (A-SPIM) system. We then suggest that the direct recording of illumination beam shapes within the working microscope system can better facilitate the analysis and subsequent re-configuration of the illumination beam to a specific geometry, and summarise the design and operation of a device that we have developed specifically for this purpose. We finally present reconstructed quantitative three dimensional flux maps of illumination beams from three microscope configurations taken using this miniature high-dynamic range beam profiling device, comparing the beam geometry of a regular SPIM system with our prototype A-SPIM system, and suggesting future improvements

Monte Carlo simulations and phantom modeling for spatial frequency domain imaging of surgical wound monitoring

Postoperative surgical wound infection is a serious problem around the globe, including in countries with advanced healthcare systems, and a method for early detection of infection is urgently required. We explore spatial frequency domain imaging (SFDI) for distinguishing changes in surgical wound healing based on the tissue scattering properties and surgical wound width measurements. A comprehensive numerical method is developed by applying a three-dimensional Monte Carlo simulation to a vertical heterogeneous wound model. The Monte Carlo simulation results are validated using resin phantom imaging experiments. We report on the SFDI lateral resolution with varying reduced scattering value and wound width and discuss the partial volume effect at the sharp vertical boundaries present in a surgical incision. The detection sensitivity of this method is dependent on spatial frequency, wound reduced scattering coefficient, and wound width. We provide guidelines for future SFDI instrument design and explanation for the expected error in SFDI measurements

Early stage dental caries detection using near infrared spatial frequency domain imaging

Early stage dental caries can be remineralized without the need for “drill-and-fill” treatments that are more invasive and less permanent. However, early stage caries lesions typically present as a white spot on a white background, resulting in many lesions only being identified after they have developed beyond the point of remineralization as cavities. We present a spatial frequency domain imaging technique to characterize the optical properties of dental tissue. This technique enables different dental tissue types (healthy enamel, healthy dentin and damaged or demineralized enamel) to be easily distinguished from one another and allows quantification of the reduced scattering coefficients of dental tissue. The use of near-infrared light at 850 nm allows high depth penetration into the tissue and suppression of absorption effects, ensuring only changes in the reduced scattering coefficient that result directly from demineralization of enamel are observed and simplifying the analysis method. This technique provides a tool to both guide the attention of dentists to areas of interest and potential demineralization, and to provide longitudinal quantified assessments to monitor caries lesion behaviour over time

Spectral index selection method for remote moisture sensing under challenging illumination conditions

Remote sensing using passive solar illumination in the Short-Wave Infrared spectrum is exposed to strong intensity variation in the spectral bands due to atmospheric changing conditions and spectral absorption. More robust spectral analysis methods, insensitive to these effects, are increasingly required to improve the accuracy of the data analysis in the field and extend the use of the system to “non ideal” illumination condition. A computational hyperspectral image analysis method (named HIAM) for deriving optimal reflectance indices for use in remote sensing of soil moisture content is detailed and demonstrated. Using histogram analysis of hyperspectral images of wet and dry soil, contrast ratios and wavelength pairings were tested to find a suitable spectral index to recover soil moisture content. Measurements of local soil samples under laboratory and field conditions have been used to demonstrate the robustness of the index to varying lighting conditions, while publicly available databases have been used to test across a selection of soil classes. In both cases, the moisture was recovered with RMS error better than 5%. As the method is independent of material type, this method has the potential to also be applied across a variety of biological and man-made samples