Donkey anti guinea pig igg biotin

Cryostat, free-floating sections of 25μm were fixed in 4% paraformaldehyde for 15 minutes and then washed extensively with PBS After incubation in 50% Formamide/2X SSC for 2h at 60°C, sections were washed again, first in 2x SSC and then in 10x PBS. After denaturation in 2N HCL at 37°C for 40 minutes, sections were made neutral by adding 0.1M Borate buffer (pH 8,5). Thereafter sections were incubated sequentially with the guinea pig anti-doublecortin (1:2000; Millipore, Germany) antibody overnight at 4°C followed by donkey anti-guinea pig IgG-biotin (Dianova, Hamburg, Germany) and streptavidin Alexa 488 (Life Technologies, Karslruhe, Germany). Finally sections were incubated with rat anti-BrdU antibody (1:2000, AbD Serotec, Puchheim, Germany). BrdU-positive cells were visualized by incubating with Cy3-conjugated donkey anti-rat IgG (H+L) (1:3000, Dianova, Hamburg, Germany).

Cryostat, free-floating sections of 25 μm were fixed in 4% paraformaldehyde for 15 min and then washed extensively with PBS. After incubation in 50% Formamide/2X SSC for 2 h at 60°C, sections were washed again, first in 2x SSC and then in 10x PBS. After denaturization in 2N HCL at 37°C for 40 min, sections were made neutral by adding 0.1 M Borate buffer (pH 8.5). Thereafter sections were incubated with the guinea pig anti-doublecortin (DCX; Millipore) antibody overnight at 4°C followed by donkey anti-guinea pig IgG-biotin (Dianova, Hamburg, Germany) and streptavidin Alexa 488 (Life Technologies, Karslruhe, Germany). Finally, sections were incubated with rat anti-BrdU antibody (1:2000, AbD Serotec, Puchheim, Germany). BrdU-positive cells were visualized by incubating with Cy3-conjugated donkey anti-rat IgG (H+L) (1:3000).