Purpose: To evaluate serum 25-hydroxyvitamin D [25(OH)D3] levels of vernal keratoconjunctivitis (VKC) children. Methods: A total of 62 non-atopic healthy children (64.5% male, mean age 10.79 ± 3.3 years) and 29 VKC children (75.9%, mean age 12.17 ± 2.7 years) were included in the study. Serum 25(OH)D3 levels measured by HPLC were compared between the two groups and a p value of <0.05 was considered as statistically significant. Results: The mean serum 25(OH)D3 level of VKC group was significantly lower than in the control group (11.02 ± 5.16 ng/mL and 15.99 ± 7.36 ng/mL, respectively) (p = 0.002). Severe vitamin D deficiency (<10 ng/mL) was detected in 48.3% of VKC children and 22.6% of the controls (p = 0.017). Time spent outdoors during daylight was higher in the control group (229.5 ± 101.2 min) compared with the VKC group (160.7 ± 65.9 min) (p = 0.008), and showed a significant correlation with serum 25(OH)D3 levels (Spearman rho = 0.812) (p<0.001). Conclusions: Children with VKC should be evaluated for vitamin D deficiency, which might occur secondary to sun avoidance.