In this study, the effect of storage at different times and temperatures on in vitro olive pollen germination was investigated. Pollens of Gemlik and Domat cultivars and GE119 and GK138 genotypes were stored at +4 °C, -20 °C, and -80 °C for 7, 35, 200, and 365 days, respectively. In vitro germination status of pollen was determined by the petri agar method, by choosing the most suitable nutrient medium for each olive. The most suitable germination media for Gemlik, Domat, GE119, and GK138 pollens were 50 ml water + 15% sucrose + 0.7% agar + 75 ppm boric acid, 50 ml water + 15% sucrose + 0.7% agar, 50 ml water + 25% sucrose + 0.5% agar, 50 ml water respectively. The interaction effect between storage time, temperatures, and cultivar on pollen germination and diameter was determined. At the end of the storage period, the highest pollen germination and diameter were observed in the Gemlik cultivar. Additionally, -80 °C temperature for Gemlik and Domat cultivars and -20 °C temperature for GE119 and GK138 genotypes were suitable for 35 days of storage. All olive pollens in the current study had germination rates below 9% in the following storage periods. The results show that storing olive pollens at sub-zero temperatures will reduce the need for daily fresh pollen collection required for important scientific studies such as breeding and artificial pollination.