Objective To validate photographic bioimaging for evaluating the severity of herpes simplex virus keratitis. Methods Stromal keratitis of patients in the Herpetic Eye Disease Study was clinically measured with a slitbeam micrometer and then photographed at trial entry. Calibrated images of 169 eyes were analyzed for the size, location, and density of stromal keratitis and endotheliitis, with shape factor as a function of area and perimeter. Validity was assessed by comparing clinical and computerized measurements and by correlating the keratitis area with visual acuity. Logistic regression explored characteristics associated with larger or denser corneal inflammation. Results Stromal keratitis had a median area of 22.4 mm2(interquartile range, 12.8-31.6 mm2) with a median shape factor of 0.69 (interquartile range, 0.56-0.79); 126 eyes (75%) had their midpoint within 2 mm of the cornea's geometric center. Photoanalytical area estimates of herpetic stromal keratitis correlated closely with clinical measurements (correlation coefficient, 0.83). Eyes with larger stromal keratitis had worse vision (correlation coefficient, 0.32) and were more likely to have iritis (P = .01). Necrotizing stromal keratitis was significantly whiter (P = .02). Conclusions Image analysis validly assesses the disciform geometry of herpetic stromal keratitis and confirms that increased severity is associated with uveitis and reduced vision.