Raman spectroscopy is a qualitative and quantitative optical technique for determining the molecular composition of matter. Improvements in spectroscopic instruments, especially the modality to detect low light level signals extended the Raman technique to biomedical applications, even in delicate structures like the eye. The purpose of this paper was to make an inventory of performed applications of Raman spectroscopy in biomedical science and especially in ophthalmology. A literature search was done using Medline, Current Contents, a patent server on the Internet, and references found in articles and patents. This search revealed a variety of Raman techniques and applications in biomedical research, and an increasing flow of articles starting in the late 1970s on Raman spectroscopy in ophthalmology. This increase in literature about Raman spectroscopy in ophthalmology feeds the expectation that this valuable technique will be introduced in the future into clinical practice.

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The purpose of this work was to obtain more quantitative knowledge about the yield of fluorescence from retinal vessels during indocyanine green angiography (ICG). The yield of fluorescence from blood was investigated for various shear rates, concentrations of ICG, and layer thicknesses. Measurements were performed in vitro on samples of human blood in a cone-plate shear chamber using frontal illumination as in scanning laser angiography. In blood and in plasma, the yield of fluorescence of ICG increased with concentration up to 0.05 and 0.1 mg/ml, respectively. At higher concentrations, the yield decreased for all layer thicknesses. For increasing layer thicknesses, both in plasma and in blood, the yield of ICG fluorescence increased nonlinearly for concentrations higher than 0.012 mg/ml. Saturation occurred for layers thicker than 200 microns in combination with ICG concentrations of 0.4 mg/ml and higher. Application of shear rates within the physiological range of the microcirculation (88/sec and 528/sec) increased the yield of fluorescence from the blood sample compared with stasis. The high transparency of blood for the excitation and emission light of ICG that was demonstrated will lead to superposition of fluorescence from superficial and deeper layers. This superposition precludes quantitative indocyanine angiography of ocular vessels.

Raman spectroscopy may offer an effective tool to non-invasively assess the local concentration of the delivered drugs within the ocular media. This technique potentially could be used to investigate the pharmacokinetics of intraocular drugs in vivo either from a releasing implant or a direct injection.