Fitc conjugated anti mouse alexa fluor 488 antibody

Brain sections were incubated with primary antibodies: anti-CD11b (1:100, abcam) or anti Iba-1 (1:100, Santa Cruz Biotechnology) in a humidified chamber at 37 °C overnight. Sections were washed with PBS and were incubated with secondary antibody FITC-conjugated anti-mouse Alexa Fluor-488 antibody (1:2000 v/v Molecular Probes, UK) for 1 h at 37 °C. Sections were laved and for nuclear staining 4′,6′-diamidino-2-phenylindole (DAPI; Hoechst, Frankfurt; Germany) 2 μg/mL in PBS was added. Sections were analysed using a Leica DM2000 microscope [51 (link)].

Brain tissue sections were incubated with one of the following primary antibodies—anti-BDNF rabbit polyclonal (1:100, Santa Cruz Biotechnology) or anti-GDNF (1:100, Santa Cruz Biotechnology)—in a humidified chamber at 37 °C overnight. Sections were washed with PBS and were incubated with secondary antibody TEXAS RED-conjugated antirabbit Alexa Fluor-594 antibody (1:1000 in PBS, v/v, Molecular Probes, Altrincham, UK) and with FITC-conjugated antimouse Alexa Fluor-488 antibody (1:2000 v/v, Molecular Probes, Altrincham, UK) for 1 h at 37 °C. Sections were laved, and for nuclear staining, 4′,6′-diamidino-2-phenylindole (DAPI; Hoechst, Frankfurt, Germany) (2 μg/mL) in PBS was added. Sections were observed and photographed at a 100× magnification using a Leica DM2000 microscope. All analyses were carried out by two observers blinded to the treatment. For immunofluorescence, a 100× magnification is shown (10-µm scale bar).

Sections were incubated with the following primary antibodies: polyclonal anti-BDNF (SCB; 1:200 in PBS, v/v), or monoclonal anti- NRF-2 (1:50; SCB) as previously described [36 (link),37 (link)]. Sections were washed with PBS and were incubated with secondary antibody TEXAS RED-conjugated anti-rabbit Alexa Fluor-594 antibody (1:1000 in PBS, v/v Molecular Probes, UK) and with FITC-conjugated anti-mouse Alexa Fluor-488 antibody (1:2000 v/v Molecular Probes, UK) for 1 h at 37 °C. Sections were rinsed and stained for nuclear signal with 4′,6′-diamidino-2-phenylindole (DAPI; Hoechst, Frankfurt; Germany) 2 μg/mL in PBS. Sections were observed and photographed using a Leica DM6 microscope (Leica Microsystems SpA, Milan, Italy).