<p>Peroxisomes are ubiquitous, oxidative organelles in eukaryotes with conserved functions in cellular lipid metabolism and redox balance. They are now recognised as key metabolic organelles of medical importance, with contributions to antiviral signalling, pathogen defence, innate immunity, cancer and neurodegeneration. Our current knowledge of peroxisomes and the molecular understanding of their diverse functions and roles in disease have been achieved through a combination of biochemical and morphological studies. This review provides a comprehensive overview of the morphological techniques and approaches developed over the past 70&#xa0;years to specifically label, identify and visualise peroxisomes in mammals, yeast and plants. We discuss the historical milestones that led to the first visualisation of peroxisomes in these organisms and trace the evolution of imaging strategies from early cytochemical and electron microscopy methods to modern fluorescence-based and high-resolution techniques. By highlighting the contribution of these approaches to advances in peroxisome biology, this review aims to place current peroxisome visualisation methods in a historical and methodological context.</p>