This article describes a method for visualizing thrombi in excised mouse brain tissue using near-infrared fluorescent imaging. The technique allows for high-contrast imaging of thromboemboli in both major cerebral arteries and small cortical blood vessels.
Begin with an excised mouse brain tissue containing thrombi, or blood clots, in the cerebral blood vessels.
The thrombi are labeled with a near-infrared fluorescent or NIRF probe that binds covalently to fibrin strands during thrombus maturation.
Place the brain tissue in a near-infrared imager, positioning the base of the brain or ventral side facing upwards to image the thrombus in the major cerebral arteries.
Apply near-infrared light to the brain tissue that excites the NIRF probe in the labeled thrombus.
Upon excitation, the probe emits light at a longer wavelength.
This causes the thrombus to fluoresce and appear bright against a dark background, minimizing interference from surrounding tissues.
Next, invert the brain tissue and image the dorsal side of the brain to visualize the thrombus in the small cortical blood vessels.
This ensures complete visualization of the thrombus in brain tissue with high contrast.
To visualize the thrombus by ex-vivo near-infrared imaging, place the excised brain tissue in the imager with the base pointing up. Obtain images of the fluorescently-labeled thromboemboli in the arteries of the Circle of Willis. Then, position the tissue with the vertex pointing up and image the cortical thromboemboli.
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