In Vivo Deep Tissue Imaging

Deep tissue imaging is a hallmark of multiphoton microscopy. The Bergamo^® III Series' ultrasensitive detector module utilizes full-field-of-view, non-descanned GaAsP PMT detectors to maximize the collection of photons scattered deep within a sample. This efficient design allows morphological features deep within a sample to be reconstructed without having to excise tissue.

The image to the left shows an XZ projection of the primary somatosensory cortex in an 8-week-old mouse expressing thy1-YFP (H line), with a sample depth in excess of 1.0 mm. The excitation wavelength was 960 nm and the data was taken with a Nikon Apo LWD 25X objective with NA 1.10. The data set is courtesy of Dr. Hajime Hirase and Katsuya Ozawa, RIKEN Brain Science Institute, Wako, Japan.

Developmental Biology

Maximum projection (dorsal and lateral, respectively) of zebrafish embryos, 48 hrs post-fertilization, showing developing neurons. Neurons are labeled with Alexa 488 against acetylated tubulin.

These images were taken using Thorlabs' confocal systems. The fluorophore was excited using a 488 nm laser and imaged with an Olympus 20X 1.0 NA water dipping lens. The data set is courtesy of Aminah Giousoh and Dr. Robert Bryson-Richardson, School of Biological Sciences, Monash University, Victoria, Australia.

Large FOV imaging at high spatiotemporal resolution is a necessary requirement for correlating neural activity of large brain volumes. This movie demonstrates the firing of neurons expressing GCaMP6f in behaving mice running on a spherical treadmill. The low magnification movie shows activity across multiple brain areas, including the somatosensory, parietal, and motor areas, at 4.3 Hz across a 6.3 mm x 5.4 mm FOV. The four non-contiguous regions were imaged at 9.6 Hz with 600 μm x 600 μm FOV, and fluorescence signals from individual neurons were sampled to cross-correlate their activity.

In these image sets, taken with the Multiphoton Mesoscope, the excitation wavelength was 970 nm. The data set is courtesy of Nicholas James Sofroniew, Daniel Flickinger, Jonathan King, and Karel Svoboda; Janelia Research Campus and Vidrio Technologies, Virginia, USA (video taken from https://elifesciences.org/content/5/e14472/article-data and used here under a Creative Commons Attribution license).

The ability to image regions within an organ of a living animal (and not just very thin tissue, like mesentery tissue) represents a major advancement in intravital microscopy. In this movie, blood perfusion can be monitored. Blood is labeled with a tail vein injection of FITC-Dextran containing red fluorescent microspheres. The microspheres can be tracked to indicate blood velocity. Red blood cells are also visible as dark shadows within the green vessels.

In these images, taken with a Thorlabs multiphoton microscope, the excitation wavelength was 780 nm and an Olympus XLUMPLFLN 20X objective with NA 1.0 was used. The preparation is courtesy of Dr. Simon Rhodes, IUPUI School of Medicine, Indianapolis, Indiana.

Deep tissue imaging is a hallmark of multiphoton microscopy. The Bergamo^® III Series' ultrasensitive detector module utilizes full-field-of-view, non-descanned GaAsP PMT detectors to maximize the collection of photons scattered deep within a sample. This efficient design allows morphological features deep within a sample to be reconstructed without having to excise tissue.

The images shown above represent an XZ projection of the primary somatosensory cortex in an 8-week-old mouse expressing thy1-YFP (H line), with a sample depth in excess of 1.0 mm. The excitation wavelength was 960 nm and the data was taken with a Nikon Apo LWD 25X objective with NA 1.10. The data set is courtesy of Dr. Hajime Hirase and Katsuya Ozawa, RIKEN Brain Science Institute, Wako, Japan.

Not all samples can be labeled with an exogenous contrast agent. Confocal reflectance microscopy allows for optical sectioning imaging by observing reflected laser light. This movie represents a Z-stack of a peachworm.

These images were taken with a Thorlabs confocal microscope. The excitation wavelength was 488 nm and the data was taken with an Olympus UPlanFl 20X objective with NA 0.50.