Human/Mouse/Rat Neurofascin Antibody

Detection of Human, Mouse, and Rat Neurofascin by Western Blot. Western blot shows lysates of rat brain tissue, mouse brain tissue, and human brain (cerebellum). PVDF Membrane was probed with 0.1 µg/mL of Chicken Anti-Human/Mouse/Rat Neurofascin Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3235) followed by HRP-conjugated Anti-Chicken IgY Secondary Antibody. Specific bands were detected for Neurofascin at approximately 140 and186 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Neurofascin in Rat Brain. Neurofascin was detected in perfusion fixed frozen sections of rat brain (dorsal root ganglion) using Chicken Anti-Human/Mouse/Rat Neurofascin Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3235) at 1.7 µg/mL overnight at 4 °C. Tissue was stained using the NorthernLights™ 557-conjugated Anti-Chicken IgY Secondary Antibody (yellow; Catalog # NL016) and counterstained with DAPI (blue). Specific staining was localized to neuronal cell bodies and Schwann cells (perinodal regions). View our protocol for Fluorescent IHC Staining of Frozen Tissue Sections.

Detection of Mouse Neurofascin by Immunohistochemistry P0T124M mutation alters SLI morphology, length,&distribution. (D) Quantification shows an increased %age of SLIs in nerves from mice harboring the P0T124M mutation (MpzT124M) at 2, 6, 12,&18 months of age. One-way ANOVA [2 month old: F (2, 10) = 29.86, p<0.0001; 6 month old: F (2, 9) = 41.60, p<0.0001, 12 month old: F (2, 6) = 34.35, p = 0.0005; 18 month old: F (2, 8) = 21.71, p = 0.0006]. Representative confocal pictures of sciatic teased fibers stained with FITC-phalloidin (ACTIN, green) (E, I,&M) or anti-MAG antibodies (Q) (green)&anti-pan-NFASC antibodies (red) at 2 (E), 6 (I),&12 (M&Q) months of age. SLI morphology is disrupted in MpzT124M mice. Scale bar: 10 μm. Measurements of SLI length at 2 (F), 6 (J),&12 (N&R) months of age. Nested one-way ANOVA [2 month old: F (2,9) = 32.16, p <0.0001; 6 month old: F (2,6) = 11.18, p = 0.0095 12 month old: F (2,9) = 6.447, p = 0.0183]. Measurements of the distance between adjacent SLI at 2 (G), 6 (K),&12 (O&S) months of age. Nested one-way ANOVA [2 month old: F (2,9) = 24.90, p = 0.0002; 6 month old: F (2,6) = 9.382, p = 0.0142; 12 month old: F (2,9) = 13.04, p = 0.0022]. Quantifications of SLI number per 100 μm at 2 (H), 6 (L),&12 (P&T) months of age. Nested one-way ANOVA [2 month old: F (2,9) = 20.29, p = 0.0005; 6 month old: F (2,6) = 22.81, p = 0.016; 12 month old: F (2,9) = 28.25, p = 0.0001]. At least 207 SLI per genotype quantified at each time point. n (animals) ≥ 3 per genotype. *p < 0.05, **p < 0.01, ***p < 0.001 by multiple-comparisons Tukey’s post hoc tests after Nested one-way ANOVA (D, F, G, H, J, K, L, R, S,&T) or by two-tailed Student’s t test (N, O,&P). Graphs indicate means ± SEMs. Image collected & cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/36350884), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Mouse Neurofascin by Immunohistochemistry P0T124M mutation alters nodes and paranodes.Representative confocal pictures of sciatic nerve teased fibers stained with antibodies against the paranodal marker CASPR (red) and the paranodal and nodal marker pan-Neurofascin (NFASC; green) at 2 (A) and 12 (E) months of age. Scale bars: 5 μm. CASPR length quantifications at 2 (B) and 12 (F) months of age. Nested one-way ANOVA [2 month old: F (2,9) = 7.009, p = 0.0146; 12 month old: F (2,7) = 35.47, p = 0.0002]. Relative frequency distributions of paranodal (CASPR) length at 2 (C) and 12 (G) months of age. Nodal length quantifications at 2 (D) and 12 (H) months of age. Nested one-way ANOVA [2 month old: F (2,9) = 1.122, p = 0.3671, 12 month old: F (2,7) = 8.012, p = 0.0155]. At least 200 paranodes and 100 nodes per genotype were quantified at each time point. n (animals) ≥ 3 per genotype. (I) Electron micrographs of ultrathin longitudinal WT and MpzT124M/T124M sciatic nerve section at 12 months of age. In (I) electron micrographs represent nodes of WT and MpzT124M/T124M sciatic nerves. (J) The quantification of nodal length shows significant widening of the node in MpzT124M/T124M. Magnifications of (I) show disorganized paranonal loops in MpzT124M/T124M but well organized in WT. n (animals) ≥ 3 per genotype, 7 to 11 nodes were counted per animals, for a total of 27 and 28 counted by genotype. Scale bare: top panel 1 μm, middle panel 500 nm, bottom panel 200 nm. ***p < 0.001 by multiple-comparisons Tukey’s post hoc tests after Nested one-way ANOVA (B, F, D, and H) or Nested two-tailed Student’s t test (J). Graphs indicate means ± SEMs. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/36350884), licensed under a CC-BY license. Not internally tested by R&D Systems.