Li-ion batteries, as a secondary battery type, are currently the most viable option for powering hybrid/electric vehicles. They have considerable advantages such as high specific energy and power, no memory effect, long cycling life, low maintenance requirement, and low self-discharge rate. However, their thermal performance can easily deteriorate at extreme ambient temperatures. Therefore, in this study, thermal and electrical behaviors of Li-ion batteries were investigated under various operating temperatures using a driving cycle that represents a highway driving condition. A battery testing system was used to determine the performance of Li-ion batteries under simulated loads. The results show that the measured temperature profiles, in broad strokes, follow the current profile. Because of the thermal capacitance of the battery, the changes in temperature variations are observed to be the smoothened out version of the current profile. Moreover, the results show that the extreme ambient temperatures have adverse effects on thermal performance of Li-ion batteries. The volumetric energy density and the capacity of the cell significantly decrease at cold ambient temperatures, especially for the sub-zero temperature applications possibly due to weak ionic conductivity within the cell. On the other hand, the difference between the ambient temperature and the surface of the cell becomes more pronounced as the ambient temperature decreases.