Objective: To investigate the changes due to wearing long-term overnight orthokeratology lenses on the stability of tear film as measured by different diagnostic devices. Methods: This was a prospective clinical study. One hundred eighteen patients, who were treated with orthokeratology lenses at The Fourth Affiliated Hospital of the China Medical University from January 2016 to October 2017, were included in the study. The patients’ ages ranged from 8 to 18 years (11.8±2.5). Measurements of tear meniscus height (TMH), noninvasive first breakup time [NIBUT (f )] and noninvasive average breakup time [NIBUT (av)] in each subject were obtained with a Keratograph 5M. Measurements were taken before lens wear and 1, 3, and 6 months after lens wear. The lipid layer interference patterns of the lacrimal film were divided by the DR-1 Tearscope Plus. Data were analyzed by repeated measures analysis of variance and by Pearson correlation analysis. Results: 1. Tear meniscus height: The TMH results showed a consistent but not statistically significant difference before and after wearing orthokeratology lenses. 2. NIBUT(f) and NIBUT(av): NIBUT had decreased by the time of the l-month visit (P<0.001); after that, all of the above changes decreased slightly and then stabilized. 3. Grades of tear film quality. There was a decrease in the stability of tear film from before lens wear to 1 month after lens wear (P<0.001), but there were no significant grade differences at 3 months and 6 months compared to prior measurements. 4. There was a small correlation between the variation in the lacrimal film lipid layer interference patterns and the BUT measured by different diagnostic devices during the follow-up study (r=-0.145, P=0.011; r= 0.147,P=0.019). Conclusions: Wearing contact lenses does not induce tear deficiency. It can result in decreasing the stability of tear film during the early stages of lens wear, but stability is gradually restored after that. Oculus keratography and DR-1 Tearscope Plus can assist in the diagnosis of dry eye diseases at different levels.