Actinides are important nuclides for the analysis of radioactive wastes arising from nuclear fuel cycle facilities. In order to achieve simple and rapid analysis of actinides, capillary electrophoresis-laser-induced fluorescent detection method (CE-LIF) is one of the potential candidates. In this study, new emissive probes of actinide ions suitable for CE-LIF were developed for the first time. The detection and separation of americium and neptunium ions as model nuclides were examined using several new emissive complexing probes, each of which possessed a fluorophore and a different chelating moiety. With a pre-capillary complexation technique without addition of the probe to separation buffer electrolyte, the highly sensitive fluorescent detection of Am and Np was successfully achieved using acyclic and macrocyclic multidentate probes. The results suggests that the probe with an acyclic hexadentate chelating moiety is suitable for detection and separation of Am and Np. The detection limit of mid–ppt levels was determined.