The role of two defects in the ambipolar character of nanocrystalline selenium poor Sb2Se3 is studied. At low temperature, the electrical transport in Sb2Se3 thin films with nm‐sized crystallites and grains is dominated by the ionization of a shallow acceptor, while at high temperature a deep donor is the leading one. The ionization energy of the deep donor agrees with theoretical reports for selenium vacancies. However, the value of the ionization energy of the shallow acceptor is in contradiction with theoretical reports pointing out to an unreported shallow defect. These findings have been confirmed by two independent experimental techniques, Electron Paramagnetic Resonance, and electrical transport as a function of temperature. The mobility of the free carriers has been found not to be limited by grain potential barriers (GPB), in contrast with polycrystalline thin films. Both findings, new shallow acceptor and zero height GPB, constitute advantages for the design of nanostructured Sb2Se3‐based solar cells and other optoelectronic devices. These results not only provide useful information for the growth of nanocrystalline Sb2Se3 thin films with ambipolar transport properties, but also about the complexity of defect physics in low‐symmetry and Q1D semiconductors.