Abstract
Abstract: :
Purpose: Several studies have measured the fluorescence that occurs during the ablation of corneal epithelium and stroma by integrating the total light received inside a solid angle onto a remote detector. The signal’s change in magnitude has been associated with the layer of tissue being removed, but the true mechanism of fluorescence is not understood due to the complexity of the ablation process. To better interpret the connection between changing signals and the nonlinear nature of ablation, a camera was used to measure the spatially distinct fluorescent response. Experiments were performed using discrete bandpass filters to evaluate the response at appropriate wavelengths. Methods: Based on existing spectral data, center wavelengths for four narrow bandpass filters were selected to capture fluorescence. An imaging system was placed on a rail perpendicular to the source of UV radiation. The laser was fired onto an enucleated porcine cornea at 10 Hz slightly off–axis. At the same time, image sequences were captured at a rate controlled by software. Once the epithelium was completely removed, water was applied to the surface, and several pulses were fired and recorded to evaluate the effect of rehydration on the fluorescent response. To account for multiple pulses in one image and improve the signal–to–noise ratio, images were averaged and filtered with Fourier techniques. Particular regions and total light responses were integrated and compared to results from earlier UV photo–detector experiments. Results: All filters provided sufficient transmission except for one, which was limited by low quantum efficiency of the CCD. Although camera settings can affect interpretation of images, the distribution of light during ablation has distinct spatial features. The brightness in each image tracks the removal of the epithelial layer. Once the epithelium is removed, intensity is constant and uniformly dim. If the stromal surface is re–wetted and ablated, a sharp brightness returns that quickly progresses outward towards an annular shape. The integrated signals from this study agree with the results of an alternative measurement technique. Conclusions: For the first time, spatial change in the fluorescent intensity distribution during ablation has been shown. Most striking is the apparent impact of water as seen in the sharp rise and drop in brightness upon ablating a fresh or re–wetted cornea. Variation in corneal hydration has been blamed for less–than–ideal outcomes in laser refractive surgery. This system will be used in tandem with measurements of ablation crater profiles in tissue to help clarify the relationship between hydration and outcome.
Keywords: cornea: basic science • imaging/image analysis: non-clinical • refractive surgery: PRK