Immunohistochemistry (PFA perfusion fixed frozen sections) - Anti-Doublecortin antibody (ab18723)
IHC image of Doublecortin staining in a section of frozen PFA perfusion fixed 6 week old rat dentate gyrus. After dissection, the tissue was further fixed in PFA overnight, washed in PBS and then incubated overnight in 30% sucrose. It was then embedded in OCT and cryosectioned. Non-specific protein-protein interactions were blocked in TBS containing 0.025% (v/v) Triton X-100, 0.3M glycine and 1% (w/v) BSA for 1h at room temperature. The section was incubated overnight at +4°C in TBS containing 0.025% (v/v) Triton X-100 and 1% (w/v) BSA with ab18723 at 1/1300 dilution and then incubated with ab150080 (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) preabsorbed, (Shown in red) 1/1000) for 1 hour at room temperature. DAPI was used to stain the cell nuclei (blue). The secondary-only control insert image is taken from an identical assay without primary antibody.
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
For IHC staining systems (automated and non-automated), customers should optimize variable parameters such as antibody concentrations and incubation times.
Immunohistochemistry (PFA perfusion fixed frozen sections) - Anti-Doublecortin antibody (ab18723)
IHC image of Doublecortin staining in a section of frozen PFA perfusion fixed 8 week old mouse SVZ. After dissection, the tissue was further fixed in PFA overnight, washed in PBS and then incubated overnight in 30% sucrose. It was then embedded in OCT and cryosectioned. Non-specific protein-protein interactions were blocked in TBS containing 0.025% (v/v) Triton X-100, 0.3M glycine and 1% (w/v) BSA for 1h at room temperature. The section was incubated overnight at +4°C in TBS containing 0.025% (v/v) Triton X-100 and 1% (w/v) BSA with ab18723 at 1/2000 dilution and then incubated with ab150080 (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) preabsorbed, (Shown in red) 1/1000) for 1 hour at room temperature. DAPI was used to stain the cell nuclei (blue). The secondary-only control insert image is taken from an identical assay without primary antibody.
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
For IHC staining systems (automated and non-automated), customers should optimize variable parameters such as antibody concentrations and incubation times.
Immunohistochemistry (PFA perfusion fixed frozen sections) - Anti-Doublecortin antibody (ab18723)
IHC image of Doublecortin staining in a section of frozen PFA perfusion fixed 8 week old mouse dentate gyrus. After dissection, the tissue was further fixed in PFA overnight, washed in PBS and then incubated overnight in 30% sucrose. It was then embedded in OCT and cryosectioned. Non-specific protein-protein interactions were blocked in TBS containing 0.025% (v/v) Triton X-100, 0.3M glycine and 1% (w/v) BSA for 1h at room temperature. The section was incubated overnight at +4°C in TBS containing 0.025% (v/v) Triton X-100 and 1% (w/v) BSA with ab18723 at 1/2000 dilution and then incubated with ab150080 (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) preabsorbed, (Shown in red) 1/1000) for 1 hour at room temperature. DAPI was used to stain the cell nuclei (blue). The secondary-only control insert image is taken from an identical assay without primary antibody.
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
For IHC staining systems (automated and non-automated), customers should optimize variable parameters such as antibody concentrations and incubation times.
Western blot - Anti-Doublecortin antibody (ab18723)
Blocking buffer: 3% Milk
Gel type: MOPS
Exposure Time: 1 minute 30 seconds
All lanes:
Western blot - Anti-Doublecortin antibody (ab18723) at 1 µg/mL
Lane 1:
Mouse brain tissue lysate - total protein (0 days) at 10 µg
Lane 2:
Rat brain tissue lysate - total protein (0 days) at 10 µg
Secondary
All lanes:
Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/50000 dilution
Predicted band size: 49 kDa
Observed band size: 45 kDa
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Doublecortin antibody (ab18723)
IHC image of ab18723 staining in Mouse 8 weeks brain formalin fixed paraffin embedded tissue section, performed on a Leica BONDTM system using the standard protocol F (with no post primary). The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution
- for 20 mins. The section was then incubated with ab18723, 0.1μg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. Secondary-only control image is shown as insert.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
