The application of UV-free electron laser photoelectron emission microscopy (UV-FEL PEEM) to measure the threshold photoelectron spectrum and photoionization potential for human eumelanosomes is described. The origin of potential artifacts and the limitations of the technique are discussed and their potential effects on the measured photoionization potential are quantified. The UV-FEL-PEEM images collected on human eumelanosomes isolated from black hair show that the organelle is photoionized by UV-B radiation. The photoionization threshold is determined to be 4.6 ± 0.2 eV. This result provides new insight into the origin of the differences between the photoionization and oxygen photoconsumption action spectra for eumelanins.