Teased Nerve Fiber and Paraffin Nerve Staining

This method was previously described.^{9 (link)} In brief, for teased nerve fibre staining, sciatic nerves were fixed in 4% paraformaldehyde (PFA) overnight and teased into individual fibres on glass slides. For paraffin nerve staining, sciatic nerves were fixed, embedded in paraffin and sectioned into 5 μm-thick slices. The slides were incubated with primary antibodies at 4°C overnight. Following a wash, the slides were incubated with secondary antibodies for 1 h. The stained slides were observed under a Leica THUNDER fluorescent microscope.

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Hu B., Moiseev D., Schena I., Faezov B., Dunbrack R., Chernoff J, & Li J. (2023). PAK2 is necessary for myelination in the peripheral nervous system. Brain, 147(5), 1809-1821.

Publication 2023

THUNDER fluorescent microscope

Corresponding Organization : Houston Methodist

Other organizations : Wayne State University, Fox Chase Cancer Center, Kazan Federal University

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Variable analysis

independent variables

Teasing of sciatic nerves into individual fibres
Fixing of sciatic nerves in 4% paraformaldehyde (PFA) overnight
Embedding of sciatic nerves in paraffin and sectioning into 5 μm-thick slices

dependent variables

Staining of teased nerve fibres or paraffin-embedded nerve sections with primary and secondary antibodies
Observation of stained slides under a Leica THUNDER fluorescent microscope

control variables

Incubation of slides with primary antibodies at 4°C overnight
Incubation of slides with secondary antibodies for 1 h

controls

No positive or negative controls were explicitly mentioned in the provided information.

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