Mouse anti gfap clone ga5

Paraffin-embedded tissue sections were deparaffinized in xylene and hydrated in gradient ethanol solutions. The sections were subjected to antigen repair by incubating them in sodium citrate buffer in a microwave for 10 minutes and then rinsed with 0.1 M phosphate-buffered saline (PBS). The sections were permeabilized with 0.3% Triton X-100 for 10 minutes and then rinsed with PBS. The sections were blocked with 5% goat serum for 1 hour at room temperature, and then incubated with primary antibody overnight at 4 ℃ in a wet box. The following primary antibodies were used: mouse anti-GFAP, clone GA5 (1:100 dilution, Millipore, Temecula, CA, USA), rabbit anti-CD68 (1:200, Abcam, Cambridge, UK), mouse anti-MBP [MBP2] (1:100, Abcam), and rabbit anti-NeuN [EPR12763] (1:200, Abcam). After three rinses in PBS for 10 minutes each, the sections were incubated with Coralite488-conjugated goat anti-rabbit IgG (1:200, Proteintech, Wuhan, China) or Cy3-conjugated goat anti-mouse IgG (1:100, Proteintech) for 1 hour at room temperature. The sections were washed in PBS 3 times and then coverslipped with a drop of anti-fade DAPI-Fluoromount G (SouthernBiotech, Birmingham, AL, USA) for nuclear counterstaining. The sections were visualized with an inverted fluorescence microscope BX51, and the images were analyzed with ImageJ software.