Assessment of body water and compartmentalization is an important experimental outcome in many studies. Whole-body impedance spectroscopy permits assessment of body water and composition; however the expense of commercially available systems can be prohibitive. PURPOSE: The purpose of this project was to develop an affordable, open-source, and precise system to perform whole-body impedance spectroscopy in rats. METHODS: Commercially validated circuit components, as described in the literature, were evaluated and compiled. An impedance spectroscopy system was designed using KiCad. Simulation and modeling of the circuit and instrumentation output was performed using LTSpice. The software was developed using Python, Arduino IDE. The system includes an internal per measurement calibration system and a constant current supply. The system was tested using serially-diluted KCl cells and an equivalent thorax circuit (RRC circuit). Validation studies were performed in anesthetized rats (2% isoflurane in 95% O2, 5% CO2) rats in-vivo (n-4, ~350 g). RESULTS: The impedance spectroscopy system was based on a built-in Arduino-Mega and AD5933 integrated circuit (Analog Devices) using open-source software. The system includes an internal per measurement calibration and constant current supply, and was able to stably iterate through excitation frequencies from 1 to 300 kHz. Admittance data obtained from rats were plotted in a complex plane. After least-squared semicircular regression analysis, there were consistent correlations within rats (triplicate) and among individual rats (n=4) on consecutive days. Total costs of the system was less than $250. CONCLUSION: We developed an affordable (open-source platform that precisely and repeatably provides whole-body impedance spectroscopy measurements in rats. The rat impedance results were qualitatively consistent with published data. This system can also be reprogrammed and reconfigured for use in other physiologic applications.