NeuroZoom™: Figure 1A shows the basic
microscope instrument setup that is controlled with NeuroZoom™
software. Up to eight standard glass slides are mounted on
the motorized microscope stage. Each slide holds 5-8 cross
sections of mouse brain tissue. The slides are automatically
analyzed for the location of every section. The sections are
then automatically imaged at very high resolution directly
into Neurome's database. Figure 1B shows the individual tiles
that are imaged as they correspond to each microscope field
of view. The tiles are reassembled automatically into a single
large-scale mosaic (Figure 1C). Finally, NeuroZoom™
aligns the sections by rotating and translating them into
3D registration to produce a single 3D dataset of the mouse
NeuroZoom™: Very high resolution images
are stored in Neurome's database by accurately controlling
microscope instrumentation and using sophisticated techniques
to stitch the images back together as a complete mosaic. The
images available for viewing in the atlas module of Neurome's
NeuroPortal™ viewing and access application ranges from
macroscopic views of the entire section, to regions of cells,
and finally to cellular and even subcellular levels of resolution.
BrainPrint™: 3D visualization of gene
expression data in a virtual mouse brain atlas using the BrainPrint™
function. Gene expression levels are correlated with specific
colors from a color scale. The Nissl-stained reference brain
used to generate the atlas is displayed in panel (B) in all
three orthogonal planes. The sagittal and horizontal planes
are virtual sections dynamically constructed from the original
coronal sections. (A) Quantitative results from in situ hybridization
analysis of mRNA expression in two different mouse strains
(left) are displayed within the basal ganglia and hippocampus
of the brain atlas. The middle panel shows the expression
levels from the DBA mouse brain and the right panel shows
the DBA / C57 ratio of expression. Scale bars on the right
show the expression levels as colors. (C-E) Display of gene
expression levels from microarray analysis in various regions
of the brain atlas. (C) Expression levels of gene X mRNA in
periventricular brain regions (BNST, hypothalamus, and PAG)
derived from three different embryonic secondary vesicles.
Scale bar on right shows signal intensity levels as colors.
(C) Specific brain regions (listed) along the rostrocaudal
neuraxis of the atlas template that demonstrate gene expression
patterns are color-coded. (D) The regional expression levels
of embryonic patterning genes based on hybridization signal
intensity in each brain region are shown in the adult mouse
brain. (Scale bar for D is in Panel B).
MRM 3D Reconstructions: 3D surface reconstructions
generated from image segmentation of the hippocampus (A),
as well as the cerebellum (B) and whole brain (C). Figure
D shows the 3D surface reconstructions within a 3D volumetric
MRM file. Note the brain is undissected and still lies within
the mouse head. Modified from Redwine et al., PNAS (2003)
Amyloid 3D reconstruction: 3D reconstruction
of amyloid beta distribution. (A) Serial coronal sections
immunostained with 3D6 were imaged and compiled into a 3D
data file. A surface reconstruction of the hippocampus is
shown in yellow. (B) Amyloid beta deposits were segmented
by thresholding and are displayed as a 3D reconstruction (red).
A surface reconstruction of the hippocampus (yellow) and a
single coronal section are shown for orientation. (C) 3D reconstruction
of amyloid beta (red) viewed from the posterior aspect of
the brain with the hippocampus shown as transparent yellow.
Note the extensive deposition in the neocortex and hippocampus,
and the central lucency representing the midbrain and caudate-putamen
with punctate amyloid beta visible in the frontal cortex and
olfactory bulb. (D) Large lakes and ribbons of amyloid beta
(cyan) were identified by automated detection of contiguous
structures within the 3D reconstruction of amyloid beta (shown
as transparent red; same angle of view as C). (E) Amyloid
beta sheets (cyan) are visible in the rostral part of the
dentate gyrus, shown against a single coronal section, with
the surface reconstruction of the hippocampus in transparent
yellow. (F) Magnified view of the amyloid beta lakes and ribbons
(cyan) in the dentate gyrus (within the transparent yellow
hippocampal surface) and extending into the retrosplenial
cortex (above). Modified from Reilly et al., PNAS (2003) 100:4837-4842.
Amyloid 3D reconstruction movie: 3D reconstruction of the amyloid deposits, shown in red. Note the extensive deposition in the neocortex and hippocampus, and the central lucency representing the midbrain and caudate-putamen with punctate amyloid visible in the frontal cortex and olfactory bulb.
Amyloid 3D reconstruction movie: Large lakes and ribbons of amyloid. These are colored blue, and were identified by automated detection of contiguous structures within the 3D reconstruction.
TOGA®: TOGA® (TOtal
Gene expression Analysis) is a patented method of identifying
and determining the concentration of nearly all of the genes
active in a sample cell or tissue. TOGA¨ automatically
identifies and simultaneously measures all genes expressed
in any cell or tissue sample and provides an inventory of
gene activity in any biological sample. The combination of
the TOGA¨ robotic assay and the integrated TOGA Portalª
informatics suite allows the researcher to assemble data from
a single experimental comparison of any gene or set of genes
across hundreds of other tissue types, time points, and conditions.
TOGA Automation: TOGA is performed on robotics as shown in pictures from the Neurome TOGA laboratory. The process proceeds clockwise: here, starting in the upper left hand corner, a liquid handling robot sets up enzymatic reactions in small 96-well plates, followed by their distribution via a robotic arm to PCR machines. The PCR amplified products in the plates are placed into capillary electrophoresis instruments shown in the lower right hand picture where fluorescent data is generated and transferred automatically to the TOGA data center shown in the bottom left hand corner.
Photoconversion Chamber: The
diOlistic method has been optimized by Neurome to label multiple
individual neurons within a single post-fixed mouse brain
slice (Wu et al, 2003). These diOlistically labeled neurons
are then photoconverted using a closed conversion chamber,
shown in Figure A, with an oxygen-enriched environment to
oxidize the fluorescently labeled neurons into a permanent
signal with sufficient signal for transmitted light brightfield
microscopy (TLBM) imaging and cell reconstruction at the higher-magnification
required for dendritic spine detection. Figure B shows confocal
laser scanning microscopy imaged dendritic spines before photoconversion.
Figure C shows the TLBM-imaged dendritic spines after photoconversion.
Note the qualitative resemblance of the individual spines
(arrows). Overall, many cells can be photoconverted in a single
pass for higher throughput. This protocol is more powerful
than the conventional methods, which are limited to the photoconversion
of only one neuron at a time. This overall protocol in use
at Neurome produces neurons with sufficient detail under TLBM
conditions to visualize dendritic spines (Wu et al., 2003).
DiOlistics: Neurome's DiOlistic
method can robustly label individual neurons in the subfields
of the hippocampal slice. After a high-throughput confocal-laser
microscopic imaging protocol (Wu et al., 2004), individual
neurons can be quickly 2-D digitized and 3-D displayed for
dendritic arbor analysis and subsequently, photoconverted
for high-magnification spine analysis.
Vaccine Technology: In situ hybridization image of intestinal Peyer's Patch probed for gene associated with M cells.