Endomicroscopy - Biomedical Optical Imaging Technologies: Design and Applications - page 260

8.2.1.4 Imaging Planes

Confocal microscopy, nonlinear microscopy, and full-field contact microscopy all
generate images in the plane oriented perpendicular to the optical axis (sometimes
termed tangential or en face ). This is different to the transverse cross section that
is conventionally used for pathology slides. However, it is possible to acquire
multiple en face images spanning a range of depths in tissue to create an “optical
stack,” amounting to an XYZ volume of data which can be manipulated to
extract a single two-dimensional image in any arbitrary plane. Optical coherence
tomography is distinct in that images are typically acquired in the XZ (cross-
sectional) plane, analogous to histopathology. The same principle of collecting
multiple 2-dimensional (XZ) images for reprocessing into a 3-D volume applies
with OCT, particularly with the recent advances in imaging speed.

8.2.1.5 Imaging Speed (Frame Rate)

Endomicroscopy systems intended for clinical use require sufficient imaging speed
to minimize the appearance of motion artifacts. The actual frame rate achieved
by a system depends on several factors, including detection sensitivity (weaker
signals require longer integration times for detection at adequate signal-to-noise
ratio), speed of the scanning mechanism for point-scanning techniques, and any
limitations on the illumination light intensity (which may be due to the available
light source brightness or due to mandated limits for safe tissue exposure). In
fluorescent techniques, light collection is the primary factor for majority of cases,
and the high NA of miniature systems is highly important to sustain practical frame
rates. Scanning mechanisms usually allow 15 frames per second or more for both
integrated scanners [ 19 ] and external scanning systems [ 20 - 22 ].
Confocal fluorescence and reflectance imaging with common, clinically
approved contrast agents (e.g., fluorescein, acetic acid) can reach 8-12 fps
[ 20 ], although endoscopic systems with miniaturized distal scanning systems are
commonly limited by the scan speed to a few or less frames per second [ 12 , 23 ].
Nonlinear microscopy generates relatively weak signals, so typically requires longer
integration times. Optical coherence tomography uses interferometric detection,
with Fourier-domain platforms offering significantly improved detection sensitivity
compared to earlier time-domain implementations. Consequently, benchtop OCT
systems have surpassed 100 frames per second [ 24 , 25 ], and endoscopic versions
are limited by the speed of the scanning mechanism, with current state-of-the-art
in the 30-60-fps range. Full-field contact imaging essentially collects light from all
points within the FOV in parallel directly on an imaging detector such as a CCD or
CMOS chip. With bright fluorophores (e.g., acriflavine, proflavine) these systems
as well as fiber-bundle-based endoscopes can stream images to an external display
at over 12 fps [ 26 ].