Purpose: To evaluate ocular comfort of lifitegrast ophthalmic solution 5.0% among patients with dry eye disease (DED) in the OPUS-3 trial. Methods: OPUS-3 was a multicenter, randomized, double-masked, placebo-controlled study. Adults with DED and recent artificial tear use were randomized 1:1 (lifitegrast:placebo) to ophthalmic drops twice daily for 84 days. On days 0 (baseline), 14, 42, and 84, drop comfort score (scale, 0–10; 0 = very comfortable, 10 = very uncomfortable) was measured at 0, 1, 2, and 3 minutes postinstillation. If the score was >3 at 3 minutes, assessment was repeated at 5, 10, and 15 minutes until score ≤3. Ocular treatment-emergent adverse events (TEAEs) were assessed. Results: Overall, 711 participants were randomized (n=357 received lifitegrast; n=354 received placebo). Drop comfort scores for lifitegrast-treated participants improved within 3 minutes of instillation (mean scores on day 84 for both study and fellow eyes: instillation: lifitegrast, 3.4, placebo, 1.0; 3 minutes: lifitegrast, 1.5, placebo, 0.7). The majority (64%–66%) of participants had scores <3 within 3 minutes postinstillation on days 14, 42, and 84. In participants with scores >3 at 3 minutes, the mean score in the lifitegrast group was similar to or better than that in the placebo group at 5, 10, or 15 minutes postinstillation. Lifitegrast appeared to be well tolerated, with ocular TEAEs rarely leading to discontinuation. Conclusion: In OPUS-3, lifitegrast appeared to be well tolerated and drop comfort scores approached placebo levels by 3 minutes postinstillation.