Since their discovery in the late 1960s, polydiacetylenes (PDA) have been in the limelight of scientific research. Upon external stimuli, such as heat, applied force, irradiation or addition of various molecules, they show unique mechanochromic characteristics, i.e. they change their colour from blue to red, and turn fluorescent. Despite the great progress, there are still many open questions in the field of polydiacetylenes: for instance, the exact mechanism of the colour change is still not clear, while the force sensing capabilities of PDA have not yet been exploited either due to the lack of quantitative analyses on their mechanochromic response. Thus, this thesis aims at addressing some of these questions and tries to help disambiguate the picture on the phase change of polydiacetylenes triggered by three different stimuli: force, addition of biomolecules and laser irradiation.