Noise equivalent spectral radiance (NESR) is a significant index representing the ultimate detection capability of infrared remote sensors. NESR calibration of highly sensitive infrared remote sensor requires an infrared radiation source with high stability, high uniformity and full field of view, and the uncertainty of its spectral radiance should be significantly lower than that of NESR of infrared remote sensor. Aiming at a new large aperture cascade integrating sphere NESR calibration system, this paper carried out experimental testing and research on the calibration uncertainty of NESR, and evaluated the influence of 11 uncertainty factors, such as the magnitude traceability of absolute spectral radiance, the uniformity and stability of integrating sphere output. The test results show that the relative uncertainty of the spectral radiance of the primary integrating sphere is better than 0.34% within the specified brightness temperature range of 303-308 K; the NESR calibration uncertainty is better than 0.1-0.0037 μW·cm^-2·sr^-1·μm^-1 within the wavelength range of 6-15 μm, which verifies the feasibility of applying the new calibration system to the NESR calibration of high-performance infrared remote sensor.