Background: Venom immunotherapy (VIT) has its effect by modulating various mediators resulting in immune tolerance. The aim of this study was to measure changes in plasma osteopontin (OPN) and serum basal tryptase (sBT) levels over the course of 1 year of VIT in children with venom allergy. Methods: Children who suffered from a large local reaction (LLR) or a systemic reaction (SR) after insect stings were included along with control subjects. Measurements were performed before the initiation of VIT and 6 and 12 months after it had been started. Results: A total of 58 children (24 with SR, 18 with LLR and 16 control subjects) with a median age of 9.5 years (range 6.7-12.8) were enrolled. The plasma OPN levels of patients with LLR [median 1,477 ng/ml, interquartile range (IQR) 1,123-1,772] were significantly higher than patients with SR (882 ng/ml, 579-1,086; p < 0.001) and healthy control subjects (1,015 ng/ml, 815-1,203; p = 0.002). A significant increase in plasma OPN levels in children was determined after the 1-year VIT. The sBT levels of children with SR (4.1 ng/ml, 3.6-5.8) were significantly higher than children with LLR (3.1 ng/ml, 2.5-4.0) and control subjects (3.0 ng/ml, 2.9-3.8; p = 0.001). There was no significant change in the sBT levels of the patients after the 1-year VIT. Conclusions: The results of our study showed higher baseline levels of OPN in children with LLR compared to control subjects and children with SR. In children with SR, OPN levels were increased after the 1-year VIT. Our results may suggest a possible association between OPN and successful VIT in children.