Ab207176

For immunofluorescence analysis, paraffin-embedded rat vagus nerve section (5 μm) were dewaxed and rehydrated, then subjected to antigen retrieval using citrate buffer (pH 9) at 95 °C for 45 min. Next, sections were incubated with a blocking solution containing 0.3% Triton X-100 for permeabilization and 3% BSA for 20 min at room temperature, and then incubated with the following primary antibodies at 4 °C overnight: Anti-alpha-synuclein (Abcam ab280377, 1:500), Anti-phospho S129 alpha-synuclein (Abcam ab51253, 1:500), Anti-S100 beta (Abcam ab52642, 1:500), Anti-S100 beta (Proteintech 66616-1-Ig, 1:500), Anti-Neurofilament heavy polypeptide (Abcam ab207176, 1:200), Anti-MJF-14 (Abcam ab209538, 1:500), Anti-TLR2 (Abcam ab209216, 1:500), Anti- interleukin-1 beta (IL-1β) (Santa cruz sc-12742, 1:50). After washing 3 times in PBS, nerve sections were incubated with corresponding secondary antibodies at 1:2000 (Abcam, Alexa Fluor conjugates) for 1 h at room temperature. DAPI (Sigma-Aldrich) was used for staining nuclei. Images were acquired with a confocal laser scanning microscopy (Zeiss LSM700, Oberkochen, Germany).

Cells or tissues were fixed in 4% paraformaldehyde for 20 min and then incubated in 0.2% Triton‐100X (Sigma‐Aldrich, USA) for 15 min. Samples were subsequently blocked with 5% BSA (Beyotime, China) overnight. The relevant primary antibody including anti‐CD206 antibody (ab64693, Abcam, USA), anti‐iNOS antibody (ab178945, Abcam, USA), anti‐F4/80 antibody (ab6640, Abcam, USA), anti‐HIF‐1α antibody (ab308433, Abcam, USA), anti‐IL‐1β antibody (ab254360, Abcam, USA), anti‐Tuj‐1 antibody (ab18207, Abcam, USA), anti‐GFAP antibody (ab7260, Abcam, USA), anti‐NFH antibody (ab207176, Abcam, USA), anti‐GAP43 antibody (ab75810, Abcam, USA), anti‐MAP2 antibody (ab5392, Abcam, USA) were then added to the samples at 4 °C overnight. Secondary antibody and DAPI were incubated with samples at 37 °C for 1 h the next day. Wash the sample with pbs three times before each operation for 5 min each time. Images were captured with a fluorescence microscope (Carl Zeiss, USA).