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Table 1 PEG-fusion protocol and rationale to immediately repair (join) cut axonal ends

From: Typical and atypical properties of peripheral nerve allografts enable novel strategies to repair segmental-loss injuries

Protocol
Steps #1–5
Completely sever and trim nerve ends Prepare nerve ends for neurorrhaphy and PEG repair/fusion
1. Priming
Solution #1
Irrigation of surgical field with hypotonic Ca2+-free saline for 1–2 min Increase axoplasmic volume. Open cut axonal ends. Expel intracellular membrane-bound vesicles/organelles
2. Protection
Solution #2
Administer 0.5–1% methylene blue (MB; an antioxidant) in distilled water for 1–2 min to opened cut ends Prevent formation of intracellular vesicles/organelles that interfere with PEG-fusion of cut ends and can seal-off each apposed cut end rather joining/fusing them
3. Co-apt cut nerve ends Perform neurorrhaphy Provide mechanical strength to epineurium to prevent PEG-fused axons from pulling apart. Closely appose cut axonal ends
4. PEG-fuse
many axons
Solution #3
Apply 50% w/w 3.35 kDa PEG in ddH2O for 1–2 min to the coaptation site Remove bound cell water to induce closely apposed, open, axonal membranes to non-specifically fuse
5. Membrane repair
Solution #4
Irrigation of coaptation site with isotonic Ca2+—containing saline Induce vesicle formation to plug/seal any axolemmal holes after PEG-induced annealing of open cut axonal ends