In the current work, a series of experiments are carried out to investigate the degradation behavior of lithium-ion batteries during overcharge cycling, as well as the influence of ambient temperature on the degradation. In which, different charge cut-off voltages (4.5, 4.8, and 5.0 V) and ambient temperatures (0°C , 20°C, 50°C, and 70°C) are included. During the overcharge process, the batteries demonstrate severe temperature rises, and several key electrochemical parameters such as the charge capacity, energy density, median voltage, and resistances all increase, revealing the deterioration of heat generation and electrode kinetics. Besides that, batteries exhibit serious degradation behavior during the overcharge cycling, which is presented through the evolution of battery temperature curves, charge voltage curves, and internal resistance curves. Moreover, the severity of degradation exacerbates with the increasing overcharge degree. Finally, it is found that deep-overcharged batteries may be more sensitive to the ambient temperature than slight-overcharged ones, where an abusive temperature can significantly aggravate the corresponding degradation.