Due to the limitations of test equipment, test methods and environmental factors, when the 50% discharge voltage of long air gap is measured in the test, the results deviate from the objective truth values. Taking the long gap discharge test of the valve-tower grading shield to the ground as an example, this paper establishes an uncertainty analysis method of U₅₀ measurement by "up-and-down method", calculates and analyzes the uncertainty of the test results from the aspects of statistical methods, high-voltage generator measurement systems, temperature and humidity changes during the test. The calculation and analysis show that the uncertainty of the test result is in the same order of magnitude as the deviation of the calculation result of the existing long air gap discharge voltage calculation model. When verifying the validity of the calculation model, it is necessary to take the uncertainty of the test value into consideration. The uncertainty introduced by the high-voltage generator measurement system is the largest. Improving the measurement and control accuracy of the high-voltage generator, and appropriately reducing the voltage difference of up-and-down method can effectively reduce the measurement uncertainty of 50% discharge voltage.