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. Author manuscript; available in PMC: 2020 Feb 15.
Published in final edited form as: J Neurosci Methods. 2018 Dec 23;314:1–12. doi: 10.1016/j.jneumeth.2018.12.015

Table 1.

PEG-fusion protocol

Step # Technique Purpose(s)
Preparation Trim nerve ends

  • Prepare nerve ends for neurorrhaphy and PEG repair/fusion
1 - Priming Irrigation of the surgical field with hypotonic Ca2+-free saline for 1-2min

  • Increase axoplasmic volume

  • Open cut axonal ends

  • Expel intracellular membrane-bound vesicles/organelles
2 - Protection Administration of 1% methylene blue (MB; an antioxidant) in distilled water for 1-2 min to the opened cut ends

  • Prevent formation of intracellular vesicles/organelles that interfere with PEG-fusion of cut ends
3 - Coapt cut peripheral nerve ends Perform neurorrhaphy

  • Provide mechanical strength to epineurium to prevent PEG-fused axons pulling apart

  • Bring and retain cut axon ends in close apposition
4 - PEG-fuse many axons Apply 50% w/w 3.35kDa PEG in distilled water for 1-2min to the coaptation site

  • Remove bound cell water to induce closely apposed, open, axonal membranes to non-specifically fuse
5 - Complete membrane repair Irrigation of the coaptation site with excess volume of isotonic Ca2+-containing saline

  • Induce vesicle formation to plug/seal any axolemmal holes after PEG-induced annealing of the open cut axonal ends