<p>This cohort study aimed to investigate the correlation between the severity of anxiety during laser in situ keratomileusis (LASIK) and fixation instability, as measured and plotted by the eye tracker during photo-ablation, and to develop a novel quantitative scale for anxiety-related fixation instability. LASIK was performed to correct myopia and hypermetropia with and without astigmatism in 2435 eyes of 2435 patients. Participants fulfilled the seven-item anxiety sub-score of the Hospital Anxiety and Depression scale questionnaire for scaling patients’ anxiety levels before LASIK into normal, borderline, and anxiety case groups. The eye-tracking pupil center position plots, patient’s heart rate, and surgeon-reported level of patient’s cooperation during the procedure were analyzed. An anxiety-related fixation instability score (FIS) was calculated, for which the best cut-off points to differentiate between normal, borderline, and anxiety case groups were defined. The FIS showed a high performance in separating participants into normal, borderline, and anxiety case groups and when used as a scale (0–90), values from 0 to 12 are considered normal, from 13 to 36 are considered borderline, and from 37 to 90 are considered anxiety cases. The FIS and scale are useful objective tools to quantify anxiety-related fixation instability during LASIK.</p>