Vascular Expansion Microscopy (VascExM): A new method for high-resolution optical imaging of the microvasculature
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Date
2020-08-07
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Johns Hopkins University
Abstract
Purpose: The resolution of optical images systems is restricted by the diffraction limit. In the past
two decades super resolution microscopy techniques have been developed to circumvent this
limitation. However, these techniques depend on state-of-the-art technological advancements. In
2015, the Boyden Lab at MIT developed a technique called expansion microscopy (ExM) which
allows nanoscale resolution imaging of tissue samples using conventional diffraction-limited
microscopes by physically expanding the specimen. Samples are embedded within polyelectrolyte
gelsthat get deprotonated in a basic environment; this cause the gel to swell in solutions like water.
The hydrogel expands, expanding the sample along with it, increasing the distance between closely
placed structures, thereby resolving them. This thesis aims to develop a tissue processing protocol,
based on ExM, for high-resolution 3D optical imaging of the vasculature in preclinical models and
to optimize this protocol across various vascular labels.
Methods: 10-100 μn sections of the brain, liver, lung, heart, and leg muscle of C57 BALB/c mice
were individually labeled with Tomato Lectin Tx-Red, Anti-Laminin Cy3 and a BriteVu and
Galbumin-Rhodamine polymer complex to compare the vasculature pre-and post-expansion.
Morphological parameters such as mean vessel diameter, area and volume were obtained by
vascular segmentation using IMARIS ®
to quantify the expansion process.
Results: Similar trends were observed post-expansion in the mean vessel diameter across the
different organs. A magnification of ~2.5x was observed in the mouse brain, leg, and liver
vasculature while a ~1.6x magnification was observed in lung vasculature. However, due to
sampling error expansion was not observed in the vasculature of mouse heart tissue samples.
Conclusion: Developed a new expansion protocol, VascExM, to obtain high resolution 3D images
of the Tomato Lectin Tx-Red labelled mouse vasculature using diffraction limited microscopes.
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Keywords
Expansion Microscopy, High Resolution Optical Imaging, Vasculature