Aim: To determine if changes in pupillary response are useful as a screening tool for diabetes and to assess whether pupillometry is associated with cardiac autonomic neuropathy. Methods: We conducted a cross-sectional study with participants drawn from two settings: a hospital and a community site. At the community site, individuals with newly diagnosed diabetes as well as a random sample of control individuals without diabetes, confirmed by oral glucose tolerance test, were selected. Participants underwent an LED light stimulus test and eight pupillometry variables were measured. Outcomes were diabetes, defined by oral glucose tolerance test, and cardiac autonomic dysfunction, determined by a positive readout on two of four diagnostic tests: heart rate response to the Valsalva manoeuvre; orthostatic hypotension; 30:15 ratio; and expiration-to-inspiration ratio. The area under the curve, best threshold, sensitivity and specificity of each pupillometry variable was calculated. Results: Data from 384 people, 213 with diabetes, were analysed. The mean (±sd) age of the people with diabetes was 58.6 (±8.2) years and in the control subjects it was 56.1 (±8.6) years. When comparing individuals with and without diabetes, the amplitude of the pupil reaction had the highest area under the curve [0.69 (sensitivity: 78%; specificity: 55%)]. Cardiac autonomic neuropathy was present in 51 of the 138 people evaluated (37.0%; 95% CI 28.8-45.1). To diagnose cardiac autonomic neuropathy, two pupillometry variables had the highest area under the curve: baseline pupil radius [area under the curve: 0.71 (sensitivity: 51%; specificity: 84%)], and amplitude of the pupil reaction [area under the curve: 0.70 (sensitivity: 82%; specificity: 55%)]. Conclusions: Pupillometry is an inexpensive technique to screen for diabetes and cardiac autonomic neuropathy, but it does not have sufficient accuracy for clinical use as a screening tool.