SCANNING SLIT CONFOCAL MICROSCOPY OF THE IN-VIVO CORNEA

The use of fixed slit and scanning slit confocal microscopes is described and illustrated for 1-D and 2-D confocal imaging of the in vivo cornea. The 1-D z scanning confocal microscope is described and its performance demonstrated with depth scans across the full thickness of the in vivo rabbit cornea. This instrument uses an axial scanning microscope objective to scan through the full thickness of the in vivo cornea, The instrument is optimized for rapid simultaneous z scans of corneal reflectance and fluorescence. The in vivo application described is the contact-lens-induced time dependence of the fluorescence intensity from the reduced pyridine nucleotides NAD(P)H located within the mitochondria of corneal epithelial cells. The depth resolution of this instrument is 6 mu m with a 100 x microscope objective and 18 mu m with a 50 x microscope objective. A second instrument based on a modified specular microscope with an internal focusing lens is described for 2-D noninvasive monitoring of corneal metabolism. Finally, a real-time, scanning slit confocal microscope is described and demonstrated for obtaining 2-D images of the in vivo human cornea. The unique imaging characteristics of this scanning slit in vivo confocal microscope is illustrated with a series of reflected light images of the normal in vivo human cornea. All of these instruments are modifications of previous instrument designs. The intellectual origins of these instruments are described.