This paper discusses how the 'Uncatchable Smile' illusion in Leonardo da Vinci's La Bella Principessa portrait was discovered. Kemp and Cotte 1 described the expression of the Princess as ambiguous and "subtle to an inexpressible degree". A combination of three methods was used (inter-observation, structured interviews, and psychophysical experiments) to identify what may underlie this 'ambiguity'. The inter-observation and the structured interview methods were firstly applied to generate experimental hypotheses that were successively tested by a series of psychophysical experiments. The combination of these research methods minimizes the impact of the researcher's beliefs and biases in the development of the research design. It emerged that the ambiguity in La Bella Principessa is triggered by a change in the perceived level of contentment in her facial expression and that this perceptual change is attributable to a visual illusion relating to her mouth. Moreover, it was found that a similar effect can be observed in the Mona Lisa. As the smile in La Bella Principessa disappears as soon as the viewer tries to 'catch it', we named this visual illusion the 'Uncatchable Smile'. The elusive quality of the Mona Lisa's smile 2 is probably why the portrait is so famous, and so the existence of a similar ambiguity in a portrait painted by Leonardo prior to the Mona Lisa is even more interesting.