THE PRESENT study was designed to display the effect of freeze- …..drying (lyophilization) technique using different freeze-drying media and storage temperatures on some parameters of buffalo bull spermatozoa. The semen samples were collected once weekly from five mature buffalo bulls maintained in Animal Reproduction Research Institute, Ministry of Agriculture, Al- Haram, Giza, Egypt. Samples were allocated into two portions; the first one was cryopreserved in Tris-Fructose-Egg yolk-Glycerol, while the second portion was immediately freeze-dried. These two portions were exposed to the same technical procedures. The freeze-dryer set was programmed at -55°C and 0.001 mbar pressure. The media tested were: EGTA solution, EDTA solution, TCM199 with Hanks salts enriched with 10% fetal calf serum (FCS) and TCM199 with Hanks salts enriched with 10% FCS and 0.2 M trehalose. The storage temperatures experienced were (4, - 20, - 80 ºC). The efficiency of each medium and storage temperature on some sperm parameters was explored. Our results revealed no motility either in raw or frozen-thawed spermatozoa after freeze-drying. In case of raw and frozen-thawed semen; with no respect to storage temperatures, the best  freeze-dried sperm parameters were observed in TCM-Trehalose medium, while the best storage temperature was (-80°C) followed by (-20°C) with no respect to the type of freeze-drying media. From the present study, it was concluded that the freeze-drying medium containing trehalose could preserve efficiently components of buffalo bull spermatozoa, especially the acrosome, while the storage temperature (-80°C) and (- 20°C) were the best for storage of freeze-dried buffalo bull spermatozoa.