Rat anti cd16 cd32

The brain sections and fixed cells were placed in 3% H₂O₂/methyl alcohol for 30 min and then placed in blocking solution for 30 min. The brain sections and cells were incubated with primary antibodies (rat anti-CD16/CD32 (1:500); BD Pharmingen, BD Biosciences; goat anti-CD206 (1:200), R&D Systems; rabbit anti-ionised calcium-binding adaptor molecule 1 (Iba-1) (1:200), Wako Chemicals, Virginia, USA) overnight at 4°C and immersed in a secondary antibody solution (Alexa Fluor 488-conjugated anti-mouse, Alexa Fluor 594-conjugated anti-goat and Alexa Fluor 594-conjugated anti-rat antibodies (1:200), Life Sciences, Paisley, UK) for 2 hours. For TUNEL (Terminal dexynucleotidyl transferase(TdT)-mediated dUTP Nick End Labeling) staining, the brain slices were incubated in working liquid (Roche) for 1 hour after blocking with 3% donkey serum. The brain sections and cells were covered with antifade medium containing DAPI (4',6-diamidino-2-phenylindole) images were acquired with a laser scanning confocal microscope (Nikon Ti-E, Japan). The number of positively stained cells per square millimetre was determined with NIH ImageJ software (Bethesda, Maryland, USA) by an investigator blinded to the experimental groups.