Resonance Raman imaging of macular pigment distributions in the human retina

We describe resonance Raman imaging (RRI) of macular pigment (MP) distributions in the living human eye. MP consists of the antioxidant carotenoid compounds lutein and zeaxanthin, is typically present in high concentrations in the healthy human macula relative to the peripheral retina, and is thought to protect this important central region from age-related macular degeneration. We demonstrate that RRI is capable of quantifying and imaging the spatially strongly varying MP distribution in the human retina. Using laser excitation of the MP molecules at 488nm, and sequential camera detection of light emitted back from the retina at the MP's strongest Raman peak position and at an off-peak position, RRI maps of MP are obtained at a resolution below 50microm within a fraction of a second per exposure. RRI imaging can be carried out with undilated pupils and provides a highly molecule-specific diagnostic imaging approach for MP distributions in human subjects.     

Bimodal spatial distribution of macular pigment: evidence of a gender relationship

The spatial distribution of the optical density of the human macular pigment measured by two-wavelength autofluorescence imaging exhibits in over half of the subjects an annulus of higher density superimposed on a central exponential-like distribution. This annulus is located at about 0.7 degrees from the fovea. Women have broader distributions than men, and they are more likely to exhibit this bimodal distribution. Maxwell's spot reported by subjects matches the measured distribution of their pigment. Evidence that the shape of the foveal depression may be gender related leads us to hypothesize that differences in macular pigment distribution are related to anatomical differences in the shape of the foveal depression.     

Multiwavelength shearography for quantitative measurements of two-dimensional strain distributions

We report on the development of a multiwavelength speckle pattern shearing interferometer for the determination of two-dimensional strain distributions. This system is based on simultaneous illumination of the object with three diode lasers that emit at different wavelengths between 800 and 850 nm. Wavelength separation and image acquisition were performed with a special optical arrangement, including narrow-bandpass filters and three black-and-white cameras. The shearographic camera with a variable shearing element, in combination with the appropriate illumination geometry, permitted us to isolate all six displacement derivatives from phase-stepped fringe patterns. The optical system and the measurement procedure were validated with two different experiments. First, the shearographic sensor head was used for the determination of in-plane displacements, and, second, in-plane strain distributions of an aluminum block caused by temperature expansion were measured.     

Optical imaging of dose distributions in Fricke gels

Ferrous-sulphate infused gels, or 'Fricke gels', encounter great interest in the field of radiation dosimetry, due to their potential for 3D radiation dose mapping. Typically, magnetic resonance (MR) relaxation rates are determined in these systems in order to derive the absorbed dose. However, when large concentration gradients are present, diffusion effects before and during the MR imaging may not be negligible. In these cases, optical techniques may represent a viable alternative. This paper describes research aimed at measuring 3D dose distributions in a Fricke-xylenol orange gel by measuring optical density with a CCD camera. This method is inexpensive and fast. A series of early experiments is described, in which optical density profiles were measured with a commercial microdensitometer for film dosimetry. The light box of the device was modified to work at 567 nm, close to the maximum absorbance of the ferric ion-xylenol orange complex. Under these conditions, the gel shows linearity with dose and high sensitivity.     

Senescence of foveal and parafoveal cone sensitivities and their relations to macular pigment density

Foveal and parafoveal increment thresholds were measured for 50 observers (12-88 years of age) under conditions that isolated retinal mechanisms dominated by short- (S-), middle- (M-), or long- (L-) wave-sensitive cones. Thresholds were obtained on the plateau of the threshold-versus-intensity function of each isolated mechanism and were referred to the retina by using individual measurements of ocular media and macular pigment density. Age-related increases in foveal thresholds, specified at the retina, were found for all three cone mechanisms. Parallel sensitivity losses for each cone mechanism were also observed at 4 degrees and 8 degrees in the temporal retina. A significant positive correlation was found between foveal macular pigment density and the S-cone, but not the M- and L-cone, log sensitivity difference (0 degrees-8 degrees) specified at the retina. This relation is expected from the hypothesis that the macular pigment protects the photoreceptors from senescent losses in sensitivity. However, because this result is independent of age, it is interpreted as being due to local gain changes resulting from differential filtering of incident light by the macular pigment between the fovea and the parafovea.     

Measurement issues in quantitative ultrasonic imaging

Ultrasonic imaging of “flesh and blood” is vulnerable to the natural variability of these media: the speed of sound is not known, not constant, and not amenable to calibration using simply shaped manufactured samples. When images of high dimensional accuracy are needed, as for image guided surgery, the “average” or “typical” values used in diagnostic ultrasound may not be good enough. In this paper, we identify the main sources of uncertainty, and we suggest and model experimental approaches to in situ calibration     

A low-energy beta-particle imaging system for measuring tritium distributions

A low-energy beta-particle imaging system was constructed for rapid measurement of the distributions of low-energy beta-emitting nuclides, especially tritium. The imaging system consists of a scintillator screen, two tapered fibers, an image intensifier, and a Polaroid film for image recording. The distortion of the observed image is less than 2.5% and the spatial resolution is more than 5 line pairs (lp)/mm. ZnS(Ag) screens with thicknesses of 5–7 mg/cm² have the highest scintillation light yield for low-energy beta-particles. The detectable flux densities for ¹⁴C and ³H were 7.3 and 80 s⁻¹ cm⁻², respectively.     

Noninterferometric quantitative phase imaging with soft x rays

We demonstrate quantitative noninterferometric x-ray phase-amplitude measurement. We present results from two experimental geometries. The first geometry uses x rays diverging from a point source to produce high-resolution holograms of submicrometer-sized objects. The measured phase of the projected image agrees with the geometrically determined phase to within +/-7%. The second geometry uses a direct imaging microscope setup that allows the formation of a magnified image with a zone-plate lens. Here a direct measure of the object phase is made and agrees with that of the magnified object to better than +/-10%. In both cases the accuracy of the phase is limited by the pixel resolution.     

Multispectral quantitative photoacoustic imaging of osteoarthritis in finger joints

We present in vivo experimental evidence that multispectral quantitative photoacoustic tomography (qPAT) has the potential to detect osteoarthritis (OA) in the finger joints. In this pilot study, two OA patients and three healthy volunteers were enrolled, and their distal interphalangeal (DIP) joints were examined photoacoustically by a multispectral PAT scanner. Images of tissue physiological/functional parameters including oxyhemoglobin, deoxyhemoglobin, oxygen saturation, and water content, along with the tissue acoustic velocity of all the examined joints, were simultaneously recovered using a finite element reconstruction algorithm for multispectral photoacoustic measurements. The recovered multispectral photoacoustic images show that the OA joints have significantly elevated water content, decreased oxygen saturation, and increased acoustic velocity compared to the normal joints.     

Uniform threshold intensity distribution-based quantitative multivariate imaging cytometry

Implementation of quantitative analytical approaches to image-based cellular assays remains a major challenge. We disclose a tool to achieve automatic rapid quantitative cellular imaging analysis based on uniform threshold intensity distribution. An acousto-optic tunable filter-based, quantitative multivariate imaging cytometer was set up to elucidate drug-induced cell death dynamics via cell viability and apoptosis/necrosis measurements in the human myeloid leukemia cell line, HL-60. Cells were treated with various drugs (camptothecin, naringenin, sodium salicylate) at different concentrations and time intervals. The developed protocol can directly depict and quantitate targeted cellular moieties, subsequently enabling a method that is applicable to various cellular assays with special reference to next generation drug discovery screening. This may also complement certain flow-cytometric measurements in studying quantitative physiology of cellular systems.     

Optical tomographic microscope for quantitative imaging of phase objects

We describe a tomographic microscope, for imaging phase objects, that makes use of the transport-of-intensity equation to estimate the phase of the transmitted light through the object. The wave-front data from optical fibers are reconstructed with an algorithm that incorporates correction for the ray bending. The reconstructed refractive-index cross sections of the fibers are found to be in agreement with the available values specified in the catalogs.     

A new methodology for quantitative LSPR biosensing and imaging

A new quantitative analysis methodology for localized surface plasmon resonance (LSPR) biosensing which determines surface-receptor fractional occupancy, as well as an LSPR imaging technique for the spatiotemporal mapping of binding events, is presented. Electron beam nanolithography was used to fabricate 20 × 20 arrays of gold nanostructures atop glass coverslips. A single biotinylated array was used to measure the association kinetics of neutravidin to the surface by spectroscopically determining the fractional occupancy as a function of time. By regenerating the same array, a reliable comparison of the kinetics could be made between control samples and neutravidin concentrations ranging from 1 μM to 50 nM. CCD-based imagery of the array, taken simultaneously with the spectroscopic measurements, reveals the binding of neutravidin to the surface as manifested by enhanced scattering over the majority of the resonance peak. The temporal resolution of the LSPR imaging technique was 200 ms and the spatial resolution was 8 μm(2).     

Scatter rejection in quantitative thermal and cold neutron imaging

The accuracy of quantitative neutron transmission radiography can be substantially decreased if highly scattering materials, such as water or plastics, exist in the sample. There are currently two main solutions to this problem: either performing experiments at a large distance between the detector and the sample or employ some numerical correction techniques. In the former case, the spatial resolution is substantially reduced by the limited beam divergence, while the latter correction requires a priori information about the sample and is limited to distances of above ∼2 cm. We demonstrate the feasibility of another technique, namely the possibility to remove the scattered neutron component from the transmitted neutron beam by a very compact polycapillary collimator. These ∼1 mm thick devices can be placed between the sample and the detector and remove most of the neutrons scattered at angles larger than the acceptance angle of the collimator (typically 1°). No image distortions above ∼10 μm scales are introduced by these collimators. The neutron transmission of highly scattering samples (water and plexiglass) is measured in our experiments with and without scatter rejection. In the latter case, the accuracy of measured transmission coefficient was substantially improved by our collimators.     

A literature review of portable fluorescence-based oil-in-water monitors

The results of a literature search on fluorescence-based portable detectors to measure the real-time concentration of oil are reported. For more than two decades, fluorometers have been commonly employed to monitor dispersed oil levels at oil spills on water. The focus of this paper has been to extract specific information from references about how the instruments were used, including set up and calibration procedures, the oil and dispersant measured, the approximate concentration range of the oil in the water column, and how the real-time data compared to traditional laboratory techniques.     

Plasma polymer coatings to aid retinal pigment epithelial growth for transplantation in the treatment of age related macular degeneration

Subretinal transplantation of functioning retinal pigment epithelial (RPE) cells grown on a synthetic substrate is a potential treatment for age-related macular degeneration (AMD), a common cause of irreversible vision loss in developed countries. Plasma polymers give the opportunity to tailor the surface chemistry of the artificial substrate whilst maintaining the bulk properties. In this study, plasma polymers with different functionalities were investigated in terms of their effect on RPE attachment and growth. Plasma polymers of acrylic acid (AC), allyl amine (AM) and allyl alcohol (AL) were fabricated and characterised using X-ray photoelectron spectroscopy (XPS) and water contact angle measurements. Octadiene (OD) hydrocarbon films and tissue culture polystyrene were used as controls. Wettability varied from hydrophobic OD to relatively hydrophilic AC. XPS demonstrated four very different surfaces with the expected functionalities. Attachment, proliferation and morphological examination of an RPE cell line and primary RPE cells were investigated. Both cell types grew on all surfaces, with the exception of OD, although the proliferation rate of primary cells was low. Good epithelial morphology was also demonstrated. Plasma polymerised films show potential as cell carrier surfaces for RPE cells in the treatment of AMD.     

High-spatial resolution mass spectrometric imaging of peptide and protein distributions on a surface

For the first time macromolecular ion microscope images have been recorded using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). Single-shot, mass-resolved images of the spatial distributions of intact peptide and protein ions over an area of 200 microm in diameter were obtained in less than 1 ms at a repetition rate of 12 Hz. The magnifying ion optics of the ion microscope allowed ion images to be obtained with a lateral resolution of 4 microm. These results prove the concept of high-resolution MALDI-MS imaging in microscope mode without the need for a tight laser focus and the accompanying sensitivity losses. The ion microscopy approach offers an improvement of several orders of magnitude in speed of acquisition compared to the conventional (microprobe) approach to MALDI-MS imaging.     

Data acquisition and postprocessing strategies for fast quantitative sodium imaging

An efficient scheme for quantitatively mapping the three-dimensional distribution of the sodium ion in vivo using magnetic resonance imaging is described. To make the methodology totally quantitative, the data acquisition scheme is performed with very short echo times and negligible T₁ saturation. Removal of signal variation due to imperfect radiofrequency (RF) response is accomplished using RF inhomogeneity maps acquired during each study. The high efficiency of the k-space trajectories allows the entire data collection process to be performed in under 10 min. The theory underlying the data collection and processing scheme is described along with representative examples acquired at 1.5 and 3.0 T. © 1997 John Wiley & Sons, Inc. Int J Imaging Syst Technol, 8, 544–550, 1997