Video-rate in vivo fluorescence imaging with a line-scanned dual-axis confocal microscope.
Video-rate optical-sectioning microscopy of living organisms would allow for the investigation of dynamic biological processes and would also reduce motion artifacts, especially for in vivo imaging applications. Previous feasibility studies, with a slow stage-scanned line-scanned dual-axis confocal (LS-DAC) microscope, have demonstrated that LS-DAC microscopy is capable of imaging tissues with subcellular resolution and high contrast at moderate depths of up to several hundred microns. However, the sensitivity and performance of a video-rate LS-DAC imaging system, with low-numerical aperture optics, have yet to be demonstrated. Here, we report on the construction and validation of a video-rate LS-DAC system that possesses sufficient sensitivity to visualize fluorescent contrast agents that are topically applied or systemically delivered in animal and human tissues. We present images of murine oral mucosa that are topically stained with methylene blue, and images of protoporphyrin IX-expressing brain tumor from glioma patients that have been administered 5-aminolevulinic acid prior to surgery. In addition, we demonstrate in vivo fluorescence imaging of red blood cells trafficking within the capillaries of a mouse ear, at frame rates of up to 30 fps. These results can serve as a benchmark for miniature in vivo microscopy devices under development.
University of Washington, Department of Mechanical Engineering, Molecular Biophotonics Laboratory, Seattle, Washington 98195, United States.
Ye
Y
Chen
Stony Brook University, Department of Biomedical Engineering, Molecular Biophotonics Laboratory, Stony Brook, New York 11794, United States.
Danni
D
Wang
Stony Brook University, Department of Biomedical Engineering, Molecular Biophotonics Laboratory, Stony Brook, New York 11794, United States.
Altaz
A
Khan
University of Washington, Department of Mechanical Engineering, Molecular Biophotonics Laboratory, Seattle, Washington 98195, United States.
Yu
Y
Wang
Barrow Brain Tumor Research Center, Division of Neurosurgical Oncology, Barrow Neurological Institute, St. Joseph's Hospital and Medical Center, Phoenix, Arizona 85013, United States.
Sabine
S
Borwege
Barrow Brain Tumor Research Center, Division of Neurosurgical Oncology, Barrow Neurological Institute, St. Joseph's Hospital and Medical Center, Phoenix, Arizona 85013, United States.
Nader
N
Sanai
University of Washington, Department of Mechanical Engineering, Molecular Biophotonics Laboratory, Seattle, Washington 98195, United States.
Jonathan T C
JT
Liu
D000818:Animals; D001932:Brain Neoplasms / Q000473:pathology*; D004867:Equipment Design; D019544:Equipment Failure Analysis; D005240:Feasibility Studies; D005910:Glioma / Q000473:pathology*; D006801:Humans; D007089:Image Enhancement / Q000295:instrumentation; D000069416:Intravital Microscopy / Q000295:instrumentation*; D018613:Microscopy, Confocal / Q000295:instrumentation*; D008856:Microscopy, Fluorescence / Q000295:instrumentation*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014741:Video Recording / Q000295:instrumentation*