The endoplasmic reticulum (ER) plays a vital role in protein folding, lipid metabolism, and calcium homeostasis, and its dysfunction is implicated in a wide range of pathologies, including cancer, neurodegenerative diseases, and metabolic disorders. Recent advances in molecular imaging and chemical biology have enabled the development of ER-targeted fluorescent probes to monitor distinct physicochemical parameters and biological processes within this organelle. This review complements existing articles by organizing ER-targeted probes primarily according to targeting strategy and structure–activity relationships, rather than by probe type or analyte alone and categorises these probes into six basic functional groups: (i) peptid linked probes, (ii) enzyme specific probes (iii) reactive oxygen species (ROS) probes, (iv) viscosity-sensitive probes, (v) probes for ER morphology and identity, (vi) biosensors developed for specific disease contexts.