Background: Pupillometry, the measurement of pupil response dynamics via the pupillary light reflex, is seldom used in the assessment of mild traumatic brain injury (mTBI). We hypothesized that there would be quantifiable differences in detailed pupil response measurements in patients with acute and chronic mTBI. Methods: We conducted 49 bilateral pupillometry measurements, in acute mTBI patients at 1-week (N=11), 2-4w (N=9), and 3-7mo post-injury (N=3); 14 patients with persistent post-traumatic symptoms (PTS) once, and healthy controls across a first visit (N=7) and second visit 2-4w later (N=5). Results: The percentage of left pupil diameter change was significantly greater in the acute mTBI group at second visit (mean=36.3% (2.96)), compared to controls at second visit (mean=31.6% (4.39)) (F=5.87, p=0.0321). We did not identify significant differences between acute mTBI patients and controls at first visit, PTS patients versus controls, and within the acute mTBI group across three longitudinal visits. Conclusion: While these preliminary data suggest that pupillometry under these conditions does not distinguish between patients who had a recent mTBI or those with PTS and healthy controls, further research is warranted investigating pupil behavior and its clinical utility in mTBI.