[Research of histochemical staining for identifying the function and morphology of fascicles in three-dimensional reconstruction of peripheral nerves].

To explore the histochemical staining for distinguishing and localizing nerve fibers and fascicles at histological level in three-dimensional reconstruction of peripheral nerves.The right median nerve was harvested from one fresh cadaver and embedded in OCT compound. The sample was serially horizontally sliced with 6 microm thickness. All sections were stained with Karnovsky-Roots method (group A, n = 30) firstly and then stained with toluidine blue (group B, n = 28) and Ponceau 2R (group C, n = 21) in proper sequence. The results of each step were taken photos (x 100). After successfully stitching, the two-dimensional panorama images were compared, including texture feature, the number and aver gray level of area showing acetylcholinesterase (AchE) activity, and result of auto microscopic medical image segmentation.In groups A, B, and C, the number of AchE-positive area was (21.63 +/- 4.06) x 10(2), (20.64 +/- 3.51) x 10(2), and (20.54 +/- 5.71) x 10(2), respectively, showing no significant difference among 3 groups (F = 0.64, P = 0.54); the mean gray level was (1.41 +/- 0.06) x 10(2), (1.10 +/- 0.05) x 10(2), and (1.14 +/- 0.07) x 10(2), respectively, showing significant differences between group A and groups B and C (P < 0.001). In the image of group A, only AchE-positive area was stained; in the image of group B, myelin sheath was obscure; and in the image of group C, axons and myelin sheath could be indentified, the character of nerve fibers could be distinguished clearly and accurately, and the image segmentation of fascicles could be achieved easier than other 2 images.The image of Karnovsky-Roots-toluidine blue-Ponceau 2R staining has no effect on the AchE-positive area in the image of Karnovsky-Roots staining and shows better texture feature. This improved histochemical process may provide ideal image for the three-dimensional reconstruction of peripheral nerves.