Clinical Relevance: Optometrists often are the first to examine previously underdiagnosed diabetic patients because of its ocular manifestations such as a change in refractive status. This study is relevant to help optometrists identify patients with diabetes using changes in pupil mydriasis as a marker. Background: This study compared the pupillary dilatation in subjects with diabetes mellitus and those without diabetes mellitus. The purpose of the study was to use this difference as a diagnostic marker for identification of diabetes in previously undiagnosed subjects who present for eye examination. Methods: An experimental study was conducted involving 40 non-diabetic and 80 diabetic subjects with or without diabetic retinopathy. Baseline pupil diameter was measured using Orbscan and Aberrometer. Mydriasis was elicited using a combination eye drop (0.8% Tropicamide and 5% Phenylephrine). Pupil diameter was measured at 15, 30, 45 and 60 minutes after the instillation of the eye drops. The difference in pupil diameter amongst the three groups was determined using factorial ANOVA to account for the effect of age. Results: There was no significant difference between the mean pupil diameter among the three groups at baseline when measured with Orbscan (p=0.187) and Aberrometer (p=0.146). There was a significant difference in the mean and mean change in pupil diameter among the three groups with either Orbscan or Aberrometer at 30, 45 and 60 minutes after the instillation of mydriatic drops. The non-diabetics have larger pupil diameter while the diabetics with retinopathy have the least pupil diameter. Conclusion: The study showed that there was a significant difference in pupil diameter between non-diabetics, diabetics without retinopathy and diabetics with retinopathy during mydriasis. Alongside a history of symptoms of diabetes, this difference could be explored as an ocular biomarker to identify previously undiagnosed diabetes in individuals who present for routine eye examination.