Abstract: Lithium-ion batteries are widely used in energy storage and new energy electric vehicles due to their superior performance, but the internal short circuit problem of lithium-ion batteries is a safety hazard during usage for energy storage and vehicle battery packs. If it cannot be detected in time, the deepening of the internal short circuit will be accompanied by an increase in heat, which will cause thermal runaway and lead to safety accidents. Diagnosing whether the battery pack has an internal short circuit and quantitatively estimating the short circuit resistance of the battery cell that has the internal short circuit can effectively prevent the occurrence of thermal runaway. This study proposes a quantitative diagnosis algorithm of Internal short circuit (ISC) based on the remaining charge capacity based on the charging curve of the lithium-ion battery module. The simulation and experimental verification of the algorithm are carried out under the conditions of different voltage acquisition accuracies, sampling periods, temperatures, and aging degrees. The results show that the proposed algorithm can accurately and quantitatively diagnose the ISC under certain conditions: (1) For serious ISC of 10 Ω level, high diagnosis accuracy can be obtained even under the conditions of 10 mV acquisition accuracy, 10 s sampling period, and variable temperature. For early ISC of 100 Ω level, the ISC resistance is smaller than the actual value and the diagnosis time is longer. To improve the accuracy and timeliness of early ISC diagnosis, the voltage acquisition accuracy, and sampling frequency should be higher than 1 mV and 1 Hz, respectively. (2) Battery aging will reduce the accuracy of ISC diagnosis, but it has little effect on the 10 Ω level ISC, and the diagnostic error of the ISC resistance is less than 6% even at an extremely low temperature (−20 ℃). The conclusions are of great significance to improve the accuracy of quantitative diagnosis of ISC for lithium-ion batteries.