The ECOSTRESS push-whisk thermal radiometer on the International Space Station provides the highest spatial resolution temperature retrievals over the ocean that are currently available. It is a precursor to the future TRISHNA (CNES/ISRO), SBG (NASA), and LSTM (ESA) 50 to 70 m scale missions. Radiance transfer simulations and triple collocations with in situ ocean observations and NOAA L2P geostationary satellite ocean temperature retrievals were used to characterize brightness temperature biases and their sources in ECOSTRESS Collection 1 (software Build 6) data for the period 12 January 2019 to 31 October 2022. Radiometric noise, non-uniformities in the focal plane array, and black body temperature dynamics were characterized in ocean scenes using L1A raw instrument data, L1B calibrated radiances, and L2 skin temperatures. The mean brightness temperature biases were −1.74, −1.45, and −1.77 K relative to radiance transfer simulations in the 8.78, 10.49, and 12.09 µm wavelength bands, respectively, and skin temperatures had a −1.07 K bias relative to in situ observations. Cross-track noise levels range from 60 to 600 mK and vary systematically along the focal plane array and as a function of wavelength band and scene temperature. Overall, radiometric uncertainty is most strongly influenced by cross-track noise levels and focal plane non-uniformity. Production of an ECOSTRESS sea surface temperature product that meets the requirements of the SST community will require calibration methods that reduce the biases, noise levels, and focal plane non-uniformities.