Raman spectroscopy for differential diagnosis of endophthalmitis and uveitis in rabbit iris in vitro

Abstract

We developed a diagnostic tool to differentiate between endophthalmitis and uveitis using Raman spectroscopy. Twenty-two New Zealand rabbits with endophthalmitis induced by Staphylococcus aureus (10 animals), noninfectious uveitis induced by lipopolysaccharide from Escherichia coli (10 animals) and controls (two animals) were analyzed. Twenty-four hours after the eyes were inoculated, iris tissue was dissected and subjected to dispersive Raman spectroscopy using an excitation source at 830 nm and a spectrograph/CCD camera to detect a Raman signal with an integration time of 50 s. With the collected spectra of endophthalmitis and uveitis, we developed a routine to classify spectra in each specimen using principal components analysis, using a leave-one-out cross-validation procedure. The mean Raman spectra of tissues with uveitis and endophthalmitis showed several bands in the region of 800e1800 cm 1, which have been attributed to nucleic acids, amino acids, proteins, and lipids. The bands at 1004, 1339, and 1555 cm 1 differed significantly (t-test, p < 0.05) between diseases. The principal components PC3 and PC4 differed significantly (ANOVA, p < 0.05) for the two tissue types, indicating that these PCs can be used to discriminate between the two diseases using Mahalanobis distance as a discriminator. This technique is useful for differentiating the spectral bands of uveitis and endophthalmitis, and the diagnostic model showed sensitivity of 89%, specificity of 100%, and accuracy of 92% using the leave-one-out cross-validation procedure. These results may be clinically relevant for differentiating endophthalmitis from uveitis, and this approach may become a noninvasive method to optimize the diagnosis of inflammatory and infectious vitreoretinal diseases.