In order to label specific facial nerve branches with a retrograde tracer, facial motor nerve and intramuscular branches of cryoanesthetized neonatal Thy1::YFP mice were visualized on an Olympus dissecting fluorescent microscope, severed with fine forceps or a tungsten needle, respectively, and 0.3 μl of lysine fixable tetramethylrhodamine-dextran (Rh-Dx; 10,000MW, Thermofisher) was applied to the proximal nerve stumps. Individual NL and OO motor pools were also labeled singly and doubly through intramuscular injection of Alexa Fluor-conjugated choleratoxin B (Alexa-CTB). Mice were perfused with 4% paraformaldehyde/0.1 M phosphate buffer (4%PFA/PB) 24 h post injection, hindbrains post-fixed in 4%PFA/PB overnight, cryoprotected in 30% sucrose/PB, and embedded in optimal cutting temperature compound (OCT, Tissue-Tek) for storage and cryosectioning. For behavioral studies, the Z/T facial nerve branches of P19 Thy1::YFP mice were visualized and severed as described above under isofluorane anesthesia.
Dissecting fluorescent microscope
Manufactured by Olympus
The Olympus Dissecting Fluorescent Microscope is a versatile instrument designed for detailed examination of biological samples. It provides high-resolution imaging by utilizing fluorescent illumination to enhance contrast and visualization of cellular structures and components.
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2 protocols using dissecting fluorescent microscope
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Retrograde Tracing of Facial Nerves
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Retrograde Tracing of Facial Nerves
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In order to label specific facial nerve branches with a retrograde tracer, facial motor nerve and intramuscular branches of cryoanesthetized neonatal Thy1::YFP mice were visualized on an Olympus dissecting fluorescent microscope, severed with fine forceps or a tungsten needle, respectively, and 0.3 μl of lysine fixable tetramethylrhodamine-dextran (Rh-Dx; 10,000MW, Thermofisher) was applied to the proximal nerve stumps. Individual NL and OO motor pools were also labeled singly and doubly through intramuscular injection of Alexa Fluor-conjugated choleratoxin B (Alexa-CTB). Mice were perfused with 4% paraformaldehyde/0.1 M phosphate buffer (4%PFA/PB) 24 h post injection, hindbrains post-fixed in 4%PFA/PB overnight, cryoprotected in 30% sucrose/PB, and embedded in optimal cutting temperature compound (OCT, Tissue-Tek) for storage and cryosectioning. For behavioral studies, the Z/T facial nerve branches of P19 Thy1::YFP mice were visualized and severed as described above under isofluorane anesthesia.
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