For layer markers, axonal markers and IUE brain analysis, embryonic mouse brains were dissected out and fixed in 4% PFA at 4°C for 6–10 h ours, washed three times with pH7.4 PBS, embedded in 3% agarose in PBS and then sectioned to 100–150 µm using a Vibratome LEICA VT1000 S (Leica). Brain sections were rinsed three time with PBS, permeabilized in 0.5% Triton X-100 in PBS for 30 min, incubated in blocking buffer (10% donkey serum, 2% BSA, 0.3% Triton X-100 in pH7.4 PBS) for 60 min. After blocking, sections were incubated with primary antibodies for SATB2 (Abcam, ab51502, 1:200), TBR1 (Abcam, ab31940, 1:400), CUX1/CDP (Santa Cruz, sc-13024, 1:1000), Ctip2 (Abcam, ab18465, 1:200), L1 (Millipore, MAB5272, 1:200), GFP (Aves Labs, GFP-1020, 1:1000) or NF165 (DSHB, 2H3, 1:120) in blocking buffer at 4°C overnight. On the next day, sections were thric e rinsed with 0.3% Triton X-100 in PBS and incubated in appropriate Alexa Fluor secondary antibodies (Life Technologies, 1:1000) in blocking buffer at 4°C overnight. On the third day, after staining with DAPI in PBS (1:500) for 30 min at room temperature, sections were washed three times with PBS and then mounted with ProLong Gold Antifade Mountant (Thermo Fisher Scientific). Images for layer makers were taken on LSM800 (Zeiss). Images for axonal markers were captured on Nikon Eclipse Ci microscope.
Ab31940
Manufactured by Santa Cruz Biotechnology
Ab31940 is a primary antibody that recognizes and binds to a specific target protein. It is designed for use in various laboratory techniques, such as immunohistochemistry and Western blotting, to detect the presence and localization of the target protein within samples.
Automatically generated - may contain errors
Lab products found in correlation
Ab51502, by Abcam (2 mentions)
Vibratome, by Leica Microsystems (2 mentions)
Bromodeoxyuridine (brdu), by Merck Group (2 mentions)
Mab3400, by Merck Group (2 mentions)
Sc 13024, by Santa Cruz Biotechnology (2 mentions)
Ab18465, by Merck Group (1 mentions)
Alexa fluor secondary antibody, by Thermo Fisher Scientific (1 mentions)
Sc 13024, by Abcam (1 mentions)
Prolong gold antifade mountant, by Thermo Fisher Scientific (1 mentions)
Leica vt1000s vibratome, by Leica camera (1 mentions)
Lsm 800, by Zeiss (1 mentions)
Eclipse ci microscope, by Nikon (1 mentions)
Ab23345, by Abcam (1 mentions)
Ab18465, by Abcam (1 mentions)
Hoechst 33258, by Thermo Fisher Scientific (1 mentions)
Vt100, by Leica camera (1 mentions)
Ab3080, by Abcam (1 mentions)
Sc 19233, by Santa Cruz Biotechnology (1 mentions)
Ab2253, by Merck Group (1 mentions)
Epr12763, by Abcam (1 mentions)
5 protocols using ab31940
1
Immunostaining of Embryonic Mouse Brain Sections
2
Quantitative Analysis of Rat Brain Development
3
Immunohistochemical Analysis of Embryonic and Postnatal Brains
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Pregnant females or postnatal animals were euthanized by lethal intreperitoneal injection of pentobarbital (50 mg/kg). Brains from embryos were dissected and fixed overnight (O.N.) in cold paraformaldehyde (PFA, 4%, pH 7.4). For postnatal brains, animals were perfused with intracardial 0.9% saline followed by cold 4% PFA. Brains were cut on a Vibratome (Leica VT100S) for immunohistochemistry (IHC). Sections were kept at 4°C in 0.1 M phosphate buffer saline (PBS) and were stained as described (Riccio et al., 2011 (link)) with the following primary antibodies: goat anti-GFP (1/500, Abcam, ab5450), goat anti-GFP (1/1000, Millipore, AB3080), rabbit anti-5-HT6R (1/500, Abcam, ab103016), rabbit anti-CUX1 (1/250, Santa Cruz, sc-13024), rabbit anti-TLE4 (1/500, Santa Cruz, sc-9125), rat anti-CTIP2 (1/500, Abcam, ab18465), rabbit anti-TBR2 (1/500, Abcam, ab23345), rabbit anti-TBR1 (1/500, Abcam, ab31940), goat anti-Ngn2 (1/50, Santa-Cruz, sc-19233), goat anti-Brn2 (1/50, Santa-Cruz, sc-6029), mouse anti-SATB2 (1/500, Abcam, ab51502), secondary goat or donkey Alexa-488, -568 and -647 antibodies (Molecular Probes, Invitrogen) raised against the appropriate species were used at a dilution of 1/1000 and sections were counterstained with Hoechst 33258 (1/10,000, Life Technologies, H3569).
4
Quantitative Analysis of Rat Brain Development
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Rat brains were fixed (E20) or perfused (P7) transcardially with chilled saline and 4% paraformaldehyde (PFA; EMS, wt/vol) and then incubated in 4% PFA overnight. Brain slices were sectioned coronally (100μm) on a vibratome (Leica microsystems). Brain slices were washed with PBS (Phosphate-buffered saline pH 7.4) and stained in PBS 0.3% Triton X-100 supplemented with 5% of donkey serum. Primary antibodies were incubated overnight at 4°C, sections were then washed with PBS and incubated in secondary antibodies for 2 hours at 22-25°C. For BrDU labeling experiments, BrDU (Sigma-Aldrich, B5002) was injected at 50mg/kg body weight intra-peritoneally 20 min prior to embryo harvest, then, brain slices were first incubated in 2N HCl for 25 min at 37°C, and then washed in PBS prior to antibody incubation. Antibodies used in this study were: Tbr2 (Millipore, AB2283), KI67 (Millipore, AB9260), Geminin (Santa-Cruz, SC-13015), Tbr1 (Abcam, ab31940), CDP (Santa-Cruz, SC-13024), NeuN (Millipore, MAB377), Cyclin D1 (ThermoScientific, RM-9104-S0), phospho-histone H3 (Abcam, ab14955), BrDU (Abcam, ab6326), Vimentin (Millipore, MAB3400).
5
Comprehensive Mouse Brain Analysis
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Mice were sacrificed at P14, P30 or 4 months by ketamine/ xylazine overdose followed by transcardial perfusion with saline solution (0.85% NaCl, 0.025% KCl, 0.02% NaHCO3, pH 6.9, 0.01% heparin, body temperature) and ice cold 4% paraformaldehyde (PFA) freshly depolymerized in 1×phosphate-buffered saline (PBS), pH 7.4. The fixed brains were carefully isolated from the skull and were further stored overnight in the same ice-cold 4% PFA solution as used for transcardial perfusion. For further storage and cryoprotection, the brains were transferred to a mixture of 20% glycerol and 2% dimethylsulfoxide in 0.1 M phosphate buffer. Consecutive horizontal sections (40 μm) were collected in six series using a freezing microtome. Corresponding brain sections from wildtype (WT) and NcaldKO/KO littermates (gender matched) were stained simultaneously for further immunohistochemical analysis as previously described (Kononenko et al., 2017 (link)). The following antibodies were used: anti-NCALD (1:100, 12925-1-AP, Proteintech), anti-NeuN (1:500, EPR 12763, Abcam), anti-TBR1 (1:500, ab31940), anti-CUX1 (1:200 sc-13024, Santa Cruz), anti-glial fibrillary acidic protein (anti-GFAP; 1:500, G3893, Sigma), anti-Ki-67 (1:500, ab15580 Abcam), anti-DCX (1:500, AB2253, Merck), anti-adenomatous polyposis coli (anti-APC; 1:500, OP80, Merck), anti-MBP (1:1,000 SMI94, Covance).
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