Mice were anaesthetised with an overdose of sodium pentobarbital and
transcardially perfused with saline followed by 4% paraformaldehyde (PFA).
Brains from pups younger than P6 were post-fixed for 4 h while brains from mice
older than P6 were post-fixed for 2 h at 4°C. Brains were sectioned
either on the sliding microtome at 30 or 40 µm as previously
described52 (link) or on a vibratome at 40
or 60 µm. All primary and secondary antibodies were diluted in PBS
containing 0.25% Triton X-100 and 2% BSA. The following antibodies were used:
goat anti-CTGF (1:200, Santa Cruz), rabbit anti cleaved-caspase-3 (1:200, Cell
Signalling), rabbit anti-dsRed (1:500, Clontech), goat anti-mCherry (1:500,
Antibodies-online), rabbit anti-GABA (1:2000, Millipore), mouse anti-GABA
(1:500, Sigma), mouse anti-NeuN (1:500, Millipore) mouse anti-parvalbumin
(1:1000, Swant), rabbit anti-parvalbumin (1:5000, Swant), rat anti-somatostatin
(1:300, Millipore) and rabbit anti-PTEN (1:500, Abcam). We used Alexa
Fluor-conjugated secondary antibodies (Invitrogen). For biotin amplification,
sections were incubated with biotinylated secondary antibody (1:200, Vector
labs), followed by Alexa Fluor-conjugated Streptavidin (1:200, Vector labs).
Blood vessels were stained with Isolectin-B4-FITC or Isolectin-B4-Dylight 594
(1:500, Vector labs).
transcardially perfused with saline followed by 4% paraformaldehyde (PFA).
Brains from pups younger than P6 were post-fixed for 4 h while brains from mice
older than P6 were post-fixed for 2 h at 4°C. Brains were sectioned
either on the sliding microtome at 30 or 40 µm as previously
described52 (link) or on a vibratome at 40
or 60 µm. All primary and secondary antibodies were diluted in PBS
containing 0.25% Triton X-100 and 2% BSA. The following antibodies were used:
goat anti-CTGF (1:200, Santa Cruz), rabbit anti cleaved-caspase-3 (1:200, Cell
Signalling), rabbit anti-dsRed (1:500, Clontech), goat anti-mCherry (1:500,
Antibodies-online), rabbit anti-GABA (1:2000, Millipore), mouse anti-GABA
(1:500, Sigma), mouse anti-NeuN (1:500, Millipore) mouse anti-parvalbumin
(1:1000, Swant), rabbit anti-parvalbumin (1:5000, Swant), rat anti-somatostatin
(1:300, Millipore) and rabbit anti-PTEN (1:500, Abcam). We used Alexa
Fluor-conjugated secondary antibodies (Invitrogen). For biotin amplification,
sections were incubated with biotinylated secondary antibody (1:200, Vector
labs), followed by Alexa Fluor-conjugated Streptavidin (1:200, Vector labs).
Blood vessels were stained with Isolectin-B4-FITC or Isolectin-B4-Dylight 594
(1:500, Vector labs).