Two-Photon Excitation Fluorescence Microscopy Motion Tracking to Study in-vivo Subcellular Structures

Project Collaborators (sorted by Institute): 
Laboratory of Cardiac Energetics, NHLBI
Project Brief: 

SPIS collaborated with the NHLBI Laboratory of Cardiac Energetics to develop a methodology enabling the study in vivo sub-cellular structures and signaling processes in real-time.  Multi-photon fluorescence imaging provides improved tissue penetration, sensitivity, and information content when investigating dynamic intercellular events within living tissue, but physiological motion degrades the quality of these images and makes temporal observations challenging.  Our system implementation, designed to be functional in tandem with the commercial two-photon microscope system, adjusts the field-of-view and focal plane of the microscope in real-time to compensate for the natural physiological motion of living tissue.  The function and performance of the commercial system is unaffected.  The motion tracking system intercepts the images as they are generated by the microscope, creating a multi-slice volume.  The system utilizes a high-performance GPU platform to correlate each real-time volume to an initial reference volume to quantify the amount and direction of tissue motion.  This motion information is used to adjust the microscope XY stage and objective to dynamically correct for the tissue motion.  SPIS integrated the custom hardware and GPU, and developed the custom software for this system. 

Two-photon excitation fluorescence microscopy motion tracking concept
Block diagram of two-photon excitation fluorescence microscopy motion tracking system
Time-lapse volume of with and without motion tracking correction