Polyimides are polymeric materials with outstanding thermal, chemical, and mechanical properties. For this reason, they find applications in several engineering sectors, including aerospace, microsystems, and biomedical applications. For realizing 3D structures made of polyimides, 3D printing is an attractive technique because it overcomes the limitations of polyimide processing using conventional manufacturing techniques such as molding and subtractive manufacturing. However, current polyimide 3D printing approaches are limited to realizing objects with the smallest dimensions of the order of a few hundred micrometers. 3D printing of polyimide objects featuring sub-micrometer resolution using two-photon polymerization by direct laser writing is demonstrated here. A negative photosensitive polyimide is applied that is widely used in microsystems applications. To demonstrate the utility of this polyimide 3D printing approach and the compatibility of the 3D objects with operation at elevated temperatures, a micro-hotplate is 3D printed and characterized at operating temperatures of above 300 °C.