Non-invasive measurement of corneal hydration

PURPOSE: To investigate the feasibility of a confocal Raman spectroscopic t echnique for the noncontact assessment of corneal hydration in vivo in two legally blind subjects. METHODS: A laser beam (632.8 nm; 15 mJ) was maintained on the cornea using a microscope objective lens (25x magnification, NA=0.5, f=10 mm) both for f ocusing the incident light as well as collecting the Raman backscattered li ght, in a 180 degrees backscatter configuration. An optical fiber, acting a s the confocal pinhole for elimination of light from out-of-focus places, w as coupled to a spectrometer that dispersed the collected light onto a sens itive array-detector for rapid spectral data acquisition over a range from 2890 to 3590 cm(-1). Raman spectra were recorded from the anterior 100 to 1 50 mum of the cornea over a period of time before and after topical applica tion of a mild dehydrating solution. The ratio between the amplitudes of th e signals at 3400 cm(-1) (OH-vibrational mode of water) and 2940 cm(-1) (CH -vibrational mode of proteins) was used as a measure of corneal hydration. RESULTS: High signal-to-noise ratio (SNR 25) Raman spectra were obtained fr om the human corneas using 15 mJ of laser light energy. Qualitative changes in the hydration of the anterior-most part of the corneas could be observe d as a result of the dehydrating agent. CONCLUSION: Confocal Raman spectroscopy could potentially be applied clinic ally as a noncontact tool for the assessment of corneal hydration in vivo.