The cave of Alistrati, is located in the Prefecture of Serres, Northern Greece near the foothills of Mount Menoikio, in the area of Petroto. This area is structured by crystalline limestones, where the development of a complex and multilevel karst system is favored. An extensive geomorphological survey was carried out for the accurate mapping of the karst surface above the cave, using UAS. For the investigation of a possible lateral extension of the existing karstic conduit, a detailed surface geophysical investigation was carried out. More specifically, three geophysical techniques were implemented: a) the Electrical Resistivity Tomography (ERT), b) the Ground Penetrating Radar (GPR) technique and c) the Very-Low Frequency (VLF) method. These 13 lines of the three geophysical techniques are fully matched at 4 locations and were join-interpreted, yielding remarkable findings. The comparative results of the above geophysical techniques, as well as their 3D presentation, highlight similar geophysical anomalies, evaluated as different types of karst system structures. Therefore, the combined geophysical survey has indicated the existence and interconnection of the first two karst levels of the area, up to a depth of 50m, as well as the possible extension of the Alistrati karstic conduit to the northeast.