Machining of micro-holes using micro-electro-discharge machining (micro-EDM) is an important field of micro-manufacturing technique. Micro-holes are essential features in optical devices, medical instruments, and automobile engine parts. Micro-EDM has been considered as the best non-traditional machining technique for generating micro-holes on high strength temperature resistant, and difficult to machine advanced materials such as Ti-6Al-4V. Ti-6Al-4V is a widely used material in aerospace and chemical industries. However, machining of micro-holes on Ti-6Al-4V using micro-EDM process is not fully explored. In this study, emphasis has been made to experimentally investigate the influence of various types of dielectrics, such as DEF-92 (EDM oil), pure deionized water, and Cu powder mixed deionized water on micro-EDM process response parameters, such as material removal rate (MRR), tool wear rate (TWR), overcut (OC), and taper during micro-hole machining of Ti-6Al-4V. It is observed that Cu powder mixed deionized water significantly improves the machining performance of the micro-EDM process. Furthermore, the machined profile and topography of the micro-hole obtained with Cu powder mixed deionized water is found to be better as compared to those machined with EDM oil or pure deionized water.