Toolleen is a town in northern Victoria, Australia. The town is on the Northern Highway and is in the Shire of Campaspe and the City of Greater Bendigo local government areas, 142 kilometres (88 mi) north of the state capital, Melbourne. At the 2011 census, Toolleen and the surrounding area had a population of 352.
During the Victorian Gold Rush and after the discovery of gold at nearby Rushworth, Toolleen was a busy intersection where the roads linking Bendigo and Beechworth and Melbourne and the Murray River port of Echuca crossed.
Toolleen Post Office opened on 12 July 1872.
The town is now mainly an agricultural region, with wine grape growing becoming more popular.