A. Neovascularization in a mouse model (with fluorescein angiography). Image taken in vivo with a Heidelberg Engineering HRA I scanning laser ophthalmoscope. Arrow points to a well-defined lesion [110].
B. Microscopic imaging of the skin of a transgenic mouse model with GFP expression in the footpad. In vivo image taken with 10-mm diameter, dual-axis confocal (DAC) microscope [111].
C. Confocal mosaic of a micronodular basal cell carcinoma (BCC) obtained ex vivo. Individual images (430 by 430-μm field of view) were combined to create a large image of excised tissue with 10- to 20-mm field of view. Small tumor nests (T) are shown as bright spots. Fluorescence emission detected from acridine orange-stained nuclei using a VivaScope 2000 by Lucid, Inc. Scale bar indicates 1 mm [112].
D. Multiphoton microcopy of intrinsic fluorescence contrast from healthy human skin at various depths. The false coloring shows the ratios of the amplitudes (a₁/a₂) of the double-exponential fluorescence lifetime decay, a presumed indicator of metabolism[6].

A. In vivo image of GFP-expressing medulloblastoma in a mouse taken by a miniature dual-axis confocal microscope designed to guide brain tumor resection (image depth is 30 μm). The design features a biaxial scanning MEMS mirror and a 2-mm diameter GRIN relay lens [46].
B. In vivo images depicting epithelial cells that have an abnormal appearance, diagnosed as serous cystadenoma. FOV is 450 microns. This image was taken with a confocal microscope designed into a laparoscopic device [117].
C. Image of a cultured mouse muscle fiber with two-photon autofluorescence (red) and second-harmonic generation (green) from sarcomeres. A GRIN relay lens was used to access subsurface muscle. Scale bar indicates 10 μm [61].

A. The spectrally-encoded scanning mechanism disperses a broadband beam into multiple beams, based on wavelength, which are projected at different angles towards the objective lens, which in-turn produces a sequence of focused points along a linear path within the specimen. A line scan is achieved rapidly in one direction by either scanning the wavelength of the laser source, or by using a broadband light source. In the latter case, the reflected light that is collected must be analyzed with a spectrograph in order to decode spatial location from the wavelength components of the signal.
B. The resonant fiber scanning mechanism uses the end of a single-mode fiber to serve as both the point source and point detector of a confocal microscope. This system can be used to gather an image by providing a mechanism for vibrating (deflecting) the end of the fiber in a resonant sinusoidal pattern to scan the focal spot within the specimen. The blue beam represents the excitation light, which is absorbed by the fluorescent molecules in the specimen. The green beam represents the fluorescence emission light collected by the end of the fiber and then directed to a detector.
C. The fiber-bundle scanning mechanism uses the ends of multiple fibers to provide multiple point sources and point detectors for a large number of parallel confocal microscopes, each imaging a single point (pixel) within a 2D image. Beam scanning is performed on the proximal end of the fiber bundle (away from the patient) where each fiber is illuminated sequentially in time.
D. A miniature dual-axis confocal microscope is depicted, where a fast electrostatically driven MEMS scanning mirror steers the excitation and emission beams along two orthogonal directions for 2D imaging. Two separate single-mode fibers act as a point source and point detector, respectively.

An example of how point-of-care pathology, via endoscopic microscopy, may be incorporated into the clinical workflow for the detection of gastrointestinal diseases. Option 1 depicts the proceduralist and pathologist working as a team. Option 2 depicts the proceduralist working by her/himself, possibly with the aid of automated image processing tools. See text for details.

A. Endomicroscopic image of a low-grade papillary tumor in a human bladder taken with a fibered confocal microscope (Mauna Kea Technologies) consisting of a 2.6 mm diameter flexible probe. Laser illumination was transmitted through an imaging bundle of more than 30,000 optical fibers. The image was taken at a depth of 60 μm with a 240-μm diameter FOV. Scale bar indicates 20 μm [113].
B. In vivo confocal fluorescence image of the border between colonic adenoma and normal mucosa, showing peptide binding to dysplastic colonocytes. Image taken in a human patient with the Cellvizio-GI imaging system by Mauna Kea Technologies, based on a fiber-bundle design [114].
C. Fibered confocal fluorescence microscopy imaging of the alveoli in the lungs of a smoker during bronchoscopy. A 1.4-mm diameter Cellvizio-Lung prototype was used (Mauna Kea Technologies), based on a fiber-bundle design. The field of view is 600 μm in diameter. Scale bar represents 50 μm [115].
D. Confocal image of the subsurface microvasculature of normal, descending human colon, collected in vivo following intravenous injection of sodium fluorescein. The confocal microscope, developed jointly by Pentax and Optiscan, is permanently integrated into an endoscope and utilizes a single fiber that is mechanically scanned with piezoelectrics. FOV = 500 × 500 microns [116].

A. The conventional confocal microscope, using a high-NA (0.8 – 1.4) objective and two pinholes to provide a point source and point detector. Images are collected by scanning the focal spot within the specimen in either reflectance or fluorescence (shown) modes. Excitation light (blue) is absorbed by the fluorescent molecules in the specimen, and fluorescence emission (green) is collected by the point detector.
B. The dual-axis confocal microscope, using two low-NA (0.1 – 0.2) objective lenses and two pinholes to provide a point source and point detector. The excitation (blue) and emission (green) beams are spatially separated within the specimen, except where they overlap at the focus.
C. The multiphoton microscope using a high-NA (0.8 – 1.4) objective and a single pinhole to provide a point source. The red beam represents the long-wavelength excitation light (high-peak-power pulsed laser), which is non-linearly absorbed by the fluorescent molecules in the specimen. The efficiency of two-photon absorption, for example, scales as the square of the intensity of the light, and is therefore confined to just the focus of the excitation beam. The green beam represents the shorter-wavelength fluorescence emission light collected by a large area detector.
D. The structured illumination microscope can be used to gather full-field (single-shot) 2D images by a photodetector array (CCD) in either reflectance or fluorescence (shown) modes. The images are quasi-optically-sectioned due to the fact that the structured pattern is only imaged with high contrast at the focal plane within the specimen. By spatially modulating the illumination pattern, out-of-focus (low-contrast) light may be distinguished from, and digitally filtered away from the in-focus light that is modulated with high contrast. The multiple blue beams represent a pattern of excitation light beams emanating from a structured light source, which is absorbed by the fluorescent molecules in the specimen. The respective green beams represent the fluorescence emission light collected by the CCD.