The papers in this special section focus on biophotonics. This is an emerging and widely growing field of advanced biomedical technologies that has opened up new horizons for extensive transfer of state-of-the-art technologies coming from the areas of quantum electronics, lasers, fiber optics and electro-optics to the life sciences and medicine. This field continues to vastly expand with advanced developments across the entire spectrum of biomedical applications ranging from fundamental laboratory bench studies to clinical patient diagnostics, bioimaging, biosensing and therapeutics. Recently developed innovative biophotonics technologies have made a significant impact on biomedical research and public health, since they provide advanced minimally invasive, cost-effective and rapid techniques for diagnostics, monitoring and treatment of a variety of diseases.

SHIGA TOXIN PATHOGENESIS IN THE GASTROINTESTINAL TRACT. Abigail J. Lenz & Andrew J. Fabich. Dept. of Biology & Chemistry, Liberty Univ., Lynchburg, VA 24515. Enterohemorrhagic Escherichia coli (EHEC) is a gram negative bacteria that is the leading cause of infectious kidney failure worldwide; a primary mechanism of EHEC infection is Shiga toxin (stx), which is carried by Golgi phosphoprotein 4 (GPP130) and binds globotriaosylceramide (Gb3) receptors on the endothelial cell surface, initiating a cascade that ultimately leads to apoptosis. Citrobacter rodentium genetically engineered with stx (CR) has successfully been used in mice to model the effects of EHEC in humans. We show that competition drives the pathogenic effects of CR infection.

NC State University, Raleigh, NC. Regulation of aflatoxin (AF) production is complex, involving transcriptional and post-transcriptional regulation focused mainly through the pathway specific transcriptional regulator aflR. An investigation into the nature of the transcriptional regulation of AF production by comparing conducive and non-conducive culture conditions revealed a clade of genes with a similar transcription profile to that of aflR. One of these genes, a putative Rho-GDP dissociation inhibitor, was characterized by gene deletion and shown to regulate AF production in Aspergillus flavus. The protein encoded by this gene, Afrdi1, showed 45% identity to Rdi1p in S. cerevisiae. The ÄAfrdi1 mutant exhibits a severe growth defect on minimal medium, a moderate growth defect on complete medium, and a temperature sensitive phenotype. Moreover, the ÄAfrdi1 mutant produces 97.3% less toxin than wild type. Inferences from S. cerevisiae reveal a possible link between AfRdi1 and RasA, which has been shown to regulate sterigmatocystin production in A. nidulans.

Some of the most commonly performed surgical operations in the world, including laser-assisted in-situ keratomileusis (LASIK), lens replacement (e.g. cataract surgery), and keratoplasty (cornea transplant), now employ therapeutic infrared femtosecond lasers (FSLs) for their extreme precision, low energy delivered into tissue and advanced ablation characteristics. Although the widely exploited applications of FSLs in medical therapeutics offer significant benefits, FSLs must generate very high intensities in order to achieve optical breakdown, the predominant tissue ablative mechanism, which can also stimulate nonlinear optical effects such as harmonic generation, an effect that generates coherent visible and UV light in the case of second- (SHG) and third-harmonic generation (THG), respectively. In order to improve the understanding of HG in corneal tissue, the effect of FSL polarization and pulse energy were investigated. FSL stimulated SHG intensity in corneal tissue was measured as the laser polarization was rotated 360 degrees. Further, the pulse energy at the SHG wavelength were measured for single FSL pulses as the pulse energy at the fundamental wavelength was varied through a range of clinically relevant values. The results of this study revealed SHG intensity oscillated with laser polarization, having a variation greater than 20%. This relationship seems to due to the intrinsic anisotropy of collagen fibril hyperpolarizability, not related to tissue birefringence. SHG pulse energy measurements showed an increase in SHG pulse energy with increasing FSL pulse energy, however conversion efficiency decreased. This may be related to the dynamic relationship between optical breakdown leading to tissue destruction and HG evolution.

PURPOSE. Optical coherence tomography (OCT) can monitor for glaucoma by measuring dimensions of the optic nerve head (ONH) cup and disc. Multiple clinical studies have shown that different OCT devices yield different estimates of retinal dimensions. We developed phantoms mimicking ONH morphology as a new way to compare ONH measurements from different clinical OCT devices.METHODS. Three phantoms were fabricated to model the ONH: One normal and two with glaucomatous anatomies. Phantoms were scanned with Stratus, RTVue, and Cirrus clinical devices, and with a laboratory OCT system as a reference. We analyzed device-reported ONH measurements of cup-to-disc ratio (CDR) and cup volume and compared them with offline measurements done manually and with a custom software algorithm, respectively.RESULTS. The mean absolute difference between clinical devices with device-reported measurements versus offline measurements was 0.082 vs. 0.013 for CDR and 0.044 mm(3) vs. 0.019 mm(3) for cup volume. Statistically significant differences between devices were present for 16 of 18 comparisons of device-reported measurements from the phantoms. Offline Cirrus measurements tended to be significantly different from those from Stratus and RTVue.CONCLUSIONS. The interdevice differences in CDR and cup volume are primarily caused by the devices' proprietary ONH analysis algorithms. The three devices yield more similar ONH measurements when a consistent offline analysis technique is applied. Scan pattern on the ONH also may be a factor in the measurement differences. This phantom-based study has provided unique insights into characteristics of OCT measurements of the ONH.

Biophotonics is an emerging field in modern biomedical technology that has opened up new horizons for transfer of state-of-the-art techniques from the areas of lasers, fiber optics and biomedical optics to the life sciences and medicine. This field continues to vastly expand with advanced developments across the entire spectrum of biomedical applications ranging from fundamental “bench” laboratory studies to clinical patient “bedside” diagnostics and therapeutics. However, in order to translate these technologies to clinical device applications, the scientific and industrial community, and FDA are facing the requirement for a thorough evaluation and review of laser radiation safety and efficacy concerns. In many cases, however, the review process is complicated due the lack of effective means and standard test methods to precisely analyze safety and effectiveness of some of the newly developed biophotonics techniques and devices. There is, therefore, an immediate public health need for new test protocols, guidance documents and standard test methods to precisely evaluate fundamental characteristics, performance quality and safety of these technologies and devices. Here, we will overview our recent developments of novel test methodologies for safety and efficacy evaluation of some emerging biophotonics technologies and medical devices. These methodologies are based on integrating the advanced features of state-of-the-art optical sensor technologies and approaches such as high-resolution fiber-optic sensing, confocal and optical coherence tomography imaging, and infrared spectroscopy. The presentation will also illustrate some methodologies developed and implemented for testing intraocular lens implants, biochemical contaminations of medical devices, ultrahigh-resolution nanoscopy, and femtosecond laser therapeutics.

Clinical therapy incorporating femtosecond laser (FSL) devices is a quickly growing field in modern biomedical technology due to their precision and ability to generate therapeutic effects with substantially less laser pulse energy. FSLs have the potential to produce nonlinear optical effects such as harmonic generation (HG), especially in tissues with significant nonlinear susceptibilities such as the cornea. HG in corneal tissue has been demonstrated in nonlinear harmonic microscopy using low-power FSLs. Furthermore, the wavelength ranges of harmonic spectral emissions generated in corneal tissues are known to be phototoxic above certain intensities. We have investigated how the critical FSL parameters pulse energy, pulse repetition rate, and numerical aperture influence both second (SHG) and third harmonic generation (THG) in corneal tissue. Experimental results demonstrated corresponding increases in HG intensity with increasing repetition rate and numerical aperture. HG duration decreased with increasing repetition rate and pulse energy. The data also demonstrated a significant difference in HG between FSL parameters representing the two most common classes of FSL therapeutic devices.

Here we present detailed transmission measurements of ex-vivo lens capsule from 40,000 cm -1 to 800 cm -1 . Time-dependent measurements were also performed to eliminate water absorption.

Purpose: The aim of this study was to evaluate the clarity of gamma-irradiated sterile corneal donor lenticules.Methods: Broadband UV, visible, and near-infrared (200-850 nm) light transmission was measured through gamma-irradiated, sterile partial-thickness, and full-thickness donor lenticules and fresh corneal tissues and compared with standard acrylic intraocular lens (IOL) implants using a conventional spectrophotometer technique.Results: All tissues had high light transmission (>= 90%) in the visible and near-infrared regions and very low (<2%) transmission below 290 nm. Differences in light transmission between irradiated and fresh cornea types were observed between 300 and 450 nm, which mirrored differences in light transmission through their respective storage solutions. Light transmission through partial-thickness irradiated donor lenticules was greatest across all wavelengths. All corneal tissues exhibited higher transmission than acrylic IOL implant across all wavelengths.Conclusions: Gamma-irradiated donor lenticules are comparable with fresh corneas regarding light transmission, with both partial-thickness and full-thickness lenticules having greater transmission than standard IOL. We would expect the optical performance of gamma-irradiated donor lenticules to be comparable to fresh cornea if used for lamellar corneal procedures that do not require a viable endothelium.

Optical coherence tomography (OCT) is a rapidly growing imaging modality, particularly in the field of ophthalmology. Accurate early diagnosis of diseases requires consistent and validated imaging performance. In contrast to more well-established medical imaging modalities, no standardized test methods currently exist for OCT quality assurance. We developed a retinal phantom which mimics the thickness and near-infrared optical properties of each anatomical retinal layer as well as the surface topography of the foveal pit. The fabrication process involves layer-by-layer spin coating of nanoparticle-embedded silicone films followed by laser micro-etching to modify the surface topography. The thickness of each layer and dimensions of the foveal pit are measured with high precision. The phantom is embedded into a commercially available, water-filled model eye to simulate ocular dispersion and emmetropic refraction, and for ease of use with clinical OCT systems. The phantom was imaged with research and clinical OCT systems to assess image quality and software accuracy. Our results indicate that this phantom may serve as a useful tool to evaluate and standardize OCT performance.

A first accurate measurement of the complex refractive index in an intralipid emulsion is demonstrated, and thereby the average scatterer particle size using standard Mie scattering calculations is extracted. Our method is based on measurement and modeling of the reflectance of a divergent laser beam from the sample surface. In the absence of any definitive reference data for the complex refractive index or particle size in highly turbid intralipid emulsions, we base our claim of accuracy on the fact that our work offers several critically important advantages over previously reported attempts. First, our measurements are in situ in the sense that they do not require any sample dilution, thus eliminating dilution errors. Second, our theoretical model does not employ any fitting parameters other than the two quantities we seek to determine, i.e., the real and imaginary parts of the refractive index, thus eliminating ambiguities arising from multiple extraneous fitting parameters. Third, we fit the entire reflectance-versus-incident-angle data curve instead of focusing on only the critical angle region, which is just a small subset of the data. Finally, despite our use of highly scattering opaque samples, our experiment uniquely satisfies a key assumption behind the Mie scattering formalism, namely, no multiple scattering occurs. Further proof of our method's validity is given by the fact that our measured particle size finds good agreement with the value obtained by dynamic light scattering.

We developed and validated a versatile test method for precise quantification of phase retardation in corneal tissues using a femtosecond laser. It is based on an experimental system for direct measurement of corneal phase rotation due to corneal birefringence effects using a dual-polarizer, computer-controlled, femtosecond laser design. It also includes a comprehensive analytical model using Jones matrices. The test method presented is used for quantification of phase retardation in corneal tissues by employing the experimental data obtained from corneal phase rotation measurements and using analytical model assessments. The experimental and theoretical results obtained, and thus, the system's high accuracy and repeatability potential for assessing the corneal phase retardation are validated using control phase retardation evaluation.

The use of femtosecond lasers (FSL) in ophthalmic procedures, such as LASIK, lens replacement (cataract surgery), as well as several other treatments, is growing rapidly. The treatment effect is based on photo ablation of ocular tissues by a series of ultra-short laser pulses. However, the laser beam characteristics change dynamically due to interactions with birefringent corneal tissue, which may affect the outcome of the laser treatment. To better understand the effect the cornea has on the laser characteristics, we developed a system for measuring retardation and validated it with precise, standard phase retarders. Then we measured the phase retardation of FSLs through bovine corneas and found that there is a considerable, location dependent, variation in retardation values. This information can potentially help optimize FSL parameters to make their application in ophthalmic procedures safer and more effective.