The study involves development of ultrasound-based regularized Nakagami imaging (RNI) that improves the quality of Nakagami image. The feasibility of RNI to image the spatial and temporal evolution of hotspot during microwave (MW) hyperthermia experiment was explored on in-vitro polyacrylamide gel (PAG)-agar based phantoms. The normalized cumulative differential regularized Nakagami (NCDRN) maps were estimated from the envelope of beamformed ultrasound radiofrequency (RF) data using proposed RNI technique. The NCDRN maps were estimated at different time instants for the entire duration of the experiment. The experiments were carried out on phantoms at power level of 12 W fed to the microwave antenna. The contour maps of the NCDRN and the ground truth temperature map, obtained using an infra-red (IR) thermal camera corresponding to ultrasound imaging plane, showed that NCDRN was able to locate the axial and lateral co-ordinates of the hotspot with an error of < 1.5 mm axially and < 0.4 mm laterally. This preliminary in-vitro study demonstrates that NCDRN maps estimated using the regularized Nakagami imaging may have potential in imaging and evaluating the spatio-temporal evolution of hotspot and may help in the development of ultrasound-based image guided hotspot monitoring system for microwave hyperthermia. © 2019 IEEE.