Objective: The purpose of this study was to investigate the safety and effectiveness of topical tacrolimus 0.05% suspension in the management of prolonged steroid-dependent and -resistant vernal keratoconjunctivitis (VKC). Materials and methods: The present interventional prospective case series were conducted on the research units suffering from refractory VKC between 2017, April and 2020, December. Despite routine drug treatment, consisting of topical steroids (steroid-resistant group), all subjects exhibited active symptomatic illness or were dependent on topical steroids (steroid-dependent group). After discontinuation of all medications for at least 1 week, the patients received eye drop (topical tacrolimus 0.05%) minimally for six months. Objective and subjective signs and symptoms were examined for alterations after one week, and one and six months of treatment. Analyses were continued for probable systemic and ocular adverse effects. Results: The study included 28 eyes of 14 participants (including 4 females and 10 males), with the mean age of 22 ± 8 years (between 11 and 38 years), and the mean VKC duration of 12 ± 6 years (between 5 and 23 years), as well as follow up period of 10±2 months (between 6 and 12 months). The results showed an improvement (P<0.001) in the symptoms such as mucus discharge, foreign body sensation, photosensitivity, redness, burning and itching. The initial sign was conjunctival hyperemia, which improved one week after treatment. Other objective signs including, conjunctival and limbal papillary hypertrophy, corneal pannus and corneal punctate epithelial erosions improved at least one month after treatment. No changes were observed in corneal opacity during follow-up (p> 0.05). There were no systemic or ocular adverse effects. Conclusion: This is the first time that tacrolimus suspension drops 0.05%, are used in these cases. Topical tacrolimus 0.05% was shown to be an effectiveness and safe agent to treat the refractory VKC. This drug is recommended to be prescribed prior to the appearance of side effects such as corneal opacity, irreversible deficiency in limbal stem cell, and steroid side effects. Keywords: Vernal keratoconjunctivitis, Tacrolimus eye drop, Refractory