Luminescent Microspheres Resolved from Strong Background on an Automated Time-Gated Luminescence Microscopy Workstation

Fluorescence microscopy is a powerful tool for the rapid identification of target organisms. However, natural autofluorescence often interferes with identification. Time-gated luminescence microscopy (TGLM) uses luminescent labels with long persistence in conjunction with digital imaging to regain discriminative power. Following the excitation pulse, short-lived autofluorescence decays rapidly whereas the long-lived emission from lanthanide doped polymer beads persists for hundreds of microseconds. After a short resolving period, a gated high gain camera captures the persistent emission in the absence of short-lived fluorescence. We report on the development of a TGLM software system for automated scanning of microscope slides, and show its use to resolve luminescent microspheres within a matrix of autofluorescent algae.