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Plast Reconstr Surg. 2018 Dec 26. doi: 10.1097/PRS.0000000000005338. [Epub ahead of print]

A Rapid Protocol for Intraoperative Assessment of Peripheral Nerve Myelinated Axon Count and its Application to Cross-Facial Nerve Grafting.



Donor nerve myelinated axon counts correlate with functional outcomes in reanimation procedures, however there exists no reliable means for their intraoperative quantification. Herein, we report a novel protocol for rapid quantification of myelinated axons from frozen sections, and demonstrate its applicability to surgical practice.


The impact of various fixation and FluoroMyelin Red™ staining strategies on resolved myelin sheath morphology from cryosections of rat and rabbit femoral and sciatic nerves was assessed. A protocol comprising fresh cryosection and rapid staining was developed, and histomorphometric results compared against conventional osmium post-fixed, resin-embedded, toluidine blue-stained sections of rat sciatic nerve. The rapid protocol was applied for intraoperative quantification of donor nerve myelinated axon count in a cross-facial nerve grafting procedure.


Resolution of myelinated axon morphology suitable for counting was realized within ten minutes of tissue harvest. Though mean myelinated axon diameter appeared larger using the rapid fresh-frozen as compared to conventional nerve processing techniques (mean ± standard deviation; rapid, 9.25 ± 0.62; conventional, 6.05 ± 0.71; p < 0.001), no difference in axon counts was observed on high power fields (rapid, 429.42 ± 49.32; conventional, 460.32 ± 69.96; p = 0.277). Whole nerve myelinated axon counts using the rapid protocol herein (8435.12 ± 1329.72) were similar to prior reports employing conventional osmium processing of rat sciatic nerve.


A rapid protocol for quantification of myelinated axon counts from peripheral nerves using widely available equipment and techniques has been described, rendering possible intraoperative assessment of donor nerve suitability for reanimation.

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