The relationship between optical properties and image contrast in confocal imaging is investigated. A Monte Carlo simulation has been developed to analyze the effects of changes in scattering, index of refraction, and absorption in a three-layer medium. Contrast was calculated from the computed signal-to-background ratios for changes in tissue optical properties. Results show that the largest source of contrast is changes in refractive index.
We present experiments to predict the maximum penetration depth atwhich typical biological structures in amelanotic tissue can bedetected with confocal microscopy. The detected signal is examinedas the signal source strength (index of refraction mismatch), thesource depth, and the medium scattering coefficient are varied. Thedetected background produced by scattering outside the focal volume isexamined as the medium scattering coefficient, the depth in the medium, the dimensionless pinhole radius, nu(p), and theshape of the scattering phase function are varied. When the systemapproaches ideal confocal performance (nu(p) ? 3), the penetration depth is limited by the signal-to-noiseratio to approximately 3-4 optical depths (OD's) for a 0.05 indexmismatch. As nu(p) increases to 8, thepenetration depth is limited by the signal-to-background ratio and isdependent on the scattering coefficient. At mu(s) = 100 cm(-1) (l(s) = 100 mum) and an index mismatch of 0.05, the maximum penetrationdepth is approximately 2 OD.
The use of high resolution, in vivo confocal imaging for noninvasive assessment of tissue pathology may offer a clinically important adjunct to standard histopathological techniques. To augment the present understanding of both the capabilities and limitations of in vivo confocal imaging, we investigated cellular sources of image contrast in amelanotic tissues, how contrast can be enhanced with external agents and how contrast is degraded by the scattering of overlying cells. A high-resolution reflected light confocal microscope was constructed and used to obtain images of various types of unstained amelanotic cells in suspension in real time before and after the addition of contrast agents. Reflectance images were compared to phase contrast images and electron micrographs to identify morphology visible with real time reflected light confocal microscopy. Mechanisms which decrease image contrast, including interference effects and scattering in overlying layers of cells, were considered. In amelanotic epithelial cells, fluctuations in the nuclear index of refraction provide signal which can be imaged even under several overlying cell layers. Acetic acid is an external contrast agent which can enhance this nuclear backscattering. Image contrast is degraded by the presence of multiple scattering in overlying cell layers. The degradation of image contrast by cell scattering depends on the scattering phase function; in vitro models which use polystyrene microspheres to approximate tissue underestimate the actual degradation caused by cell scattering. The loss in contrast can be explained using a finite difference time domain model of cellular scattering. We conclude that near real time reflected light confocal microscopy can be used to study cell morphology in vivo. Contrast degradation due to overlying tissue is a concern and cannot adequately be modeled using conventional tissue phantoms; however, acetic acid may be used to substantially increase intrinsic contrast, allowing imaging at significant depths despite distortion from overlying layers. © 1998 Society of Photo-Optical Instrumentation Engineers.
In this study, the effectiveness of pulsed and continuous wave (CW) holmium: YAG lasers in coagulating in vitro pig corneas was analyzed. With the CW laser, irradiance and exposure time were varied; irradiance, from 162 to 324 W/cm2 and exposure time, from 200 to 800 ms. With the pulsed laser, number of pulses and radiant exposure were varied; number of pulses per lesion, from 4 to 30 and radiant exposure, from 10 to 25 J/cm2. Laser-induced corneal damage was determined by analyzing histological cross sections of each lesion. Depth and diameter of the lesions were plotted against the varying laser parameters. Light and birefringent photomicrographs of typical lesion histology show that the pulsed laser significantly damaged superficial layers of the cornea and could not achieve the coagulation depths produced by the CW laser. Additional histology demonstrates that minimal surface damage (intrastromal coagulation) occurred when the CW laser beam was delivered with a sapphire-tipped contact probe. The results provide empirical data on the sensitivity of each parameter in producing a range of coagulation end points. In addition, the experimental results describe trends between the parameters of either laser and the extent of coagulation.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.