Ab18465

Manufactured by Abcam

Sourced in United Kingdom, United States, Germany

Ab18465 is a laboratory equipment product offered by Abcam. It is a device designed for use in scientific research and analysis. The core function of this product is to perform a specific task or measurement within a laboratory setting. No further details or interpretations about the intended use or applications of this product are provided.

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Lab products found in correlation

Ab31940, by Abcam (24 mentions) Ab51502, by Abcam (15 mentions) Ab23345, by Abcam (12 mentions) Mab377, by Merck Group (7 mentions) Ab13970, by Abcam (7 mentions) Sc 13024, by Abcam (6 mentions) Ab6326, by Abcam (6 mentions) Bovine serum albumin (bsa), by Merck Group (5 mentions) Sc 13024, by Santa Cruz Biotechnology (5 mentions) 4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (5 mentions) Prolong gold antifade mountant, by Thermo Fisher Scientific (4 mentions) A2052, by Merck Group (4 mentions) Ab19857, by Abcam (4 mentions) Ab97959, by Abcam (4 mentions) Ab144p, by Merck Group (4 mentions) Ab17721, by Abcam (4 mentions) Ne per nuclear and cytoplasmic extraction reagent, by Thermo Fisher Scientific (4 mentions) Hoechst 33258, by Thermo Fisher Scientific (3 mentions) T8660, by Merck Group (3 mentions) Mab354, by Merck Group (3 mentions)

Spelling variants of this product

Ctip2 ab18465 (3 citations)

94 protocols using ab18465

Immunostaining of Neuronal Markers

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Cells were washed with PBS followed by fixation with 4% paraformaldehyde (Merck Millipore) and subsequent washing. Fixed cells were permeabilized and blocked with 5% bovine serum albumin (Sigma-Aldrich) in PBS with 0.1% Triton X-100 (Carl Roth). Afterwards, cells were stained with anti-β-III-tubulin (TUJ, mouse, 1:1000, T8660, Sigma-Aldrich), anti-CTIP2 (rat, 1:200, ab18465, Abcam), anti-HSF1 (rabbit, 1:500, 4356T, Cell Signaling Technology) and/or anti-HSP70 (mouse, 1:500, ADI-SPA-810, Enzo Life Sciences), all followed by labeling with Alexa Fluor-conjugated secondary antibodies (Invitrogen). Hoechst 33258 staining (1:10,000, H1398, Invitrogen) was used to counterstain for nuclei. Coverslips were mounted with Dako mounting solution (Agilent Dako) onto microscope slides. Four or five random fields per coverslip per cell line were used for quantification. Images were acquired using an Observer Z1 fluorescence microscope (Zeiss), and exposure time was kept constant. Quantifications were conducted using the cell counter plug-in in FiJi (Schindelin et al., 2012 (link)) and threshold mask settings in ImageJ (https://imagej.nih.gov/ij/).

Schuster S., Heuten E., Velic A., Admard J., Synofzik M., Ossowski S., Macek B., Hauser S, & Schöls L. (2020). CHIP mutations affect the heat shock response differently in human fibroblasts and iPSC-derived neurons. Disease Models & Mechanisms, 13(10), dmm045096.

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Antibody Characterization for Cell Biology Research

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The following antibodies were used in this study: γ-tubulin (aa 38–53) GTU-88 (T6557, mouse, used at 1/10000 for WB and 1/1000 for IF/IHC, Sigma); Gamma-tubulin (434–451) TU-30 (ab27074, mouse, used at 1/1000, Abcam); GAPDH (MAB374, mouse, used at 1/1000, Chemicon); Cux1 (sc-13024, rabbit, used at 1/100, Santa Cruz); Cux1 (11733-AP, used at 1/100, Proteintech); GFP (A10262, chicken, used at 1/1000, ThermoFisher); CTIP2 (ab18465, rat, used at 1/500, Abcam); NeuN (MAB377, mouse, used at 1/500, Millipore); SATB2 (ab51502, mouse, used at 1/400, Abcam); TBR1 (ab31940, rabbit, used at 1/500, Abcam); Pax6 (PRB-278P, rabbit, used at 1/100, Eurogentec); Tbr2 (14–4875–80, rat, used at 1/200, eBioscience); PH3 (06–570, rabbit, used at 1/500, Millipore); Ki67 (NCL-L-Ki67-MM1, mouse, used at 1/500, Leica); Anti-tRFP (AB234, rabbit used at 1/5000 for WB, Evrogen); GCP4 (sc-271876, mouse, used at 1/1000 for WB, Santa Cruz); Anti-mouse antibody conjugated with HRP (W402B, goat, used at1/10000 for WB, Promega); Anti-rabbit antibody conjugated with HRP (W401B, goat, used at 1/100000 for WB, Promega). Anti-GCP2 antibody GCP2-01 (mouse monoclonal IgG2b) used for immunoprecipitation was described previously^{58 (link)}; Anti-GCP2 antibody GCP2-02 (mouse monoclonal IgG1, in the form of hybridoma spent culture supernatant, used at 1/10 for WB) was described previously^{58 (link)}.

Ivanova E.L., Gilet J.G., Sulimenko V., Duchon A., Rudolf G., Runge K., Collins S.C., Asselin L., Broix L., Drouot N., Tilly P., Nusbaum P., Vincent A., Magnant W., Skory V., Birling M.C., Pavlovic G., Godin J.D., Yalcin B., Hérault Y., Dráber P., Chelly J, & Hinckelmann M.V. (2019). TUBG1 missense variants underlying cortical malformations disrupt neuronal locomotion and microtubule dynamics but not neurogenesis. Nature Communications, 10, 2129.

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Analyzing Layer-Specific Neuronal Expression

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Intron-RED and pre-miR-128-2-RED electroporated pups from the same litter were analyzed at P0 (born E20). 10 µm cryosections were stained for dsRed (Abcam ab62341 1:150), Cux1 (Santa Cruz Biotechnology, sc-13024 1:150, Heidelberg, Germany), and Ctip2 (Abcam ab18465 1:500). The slices were imaged using a Leica SL confocal microscope with 40× objective. Using the Cell Counter plugin for Fiji both the total number of electroporated neurons in the cortical plate and the number of electroporated neurons positive for either Cux1 or Ctip2 was counted. The number of neurons positive for the layer marker was normalized to the total number of electroporated neurons. Three independent brains electroporated with pre-miR-128-2-RED and one brain electroporated with Intron-RED were analyzed and for each layer marker at least three slices per brain were counted.

Franzoni E., Booker S.A., Parthasarathy S., Rehfeld F., Grosser S., Srivatsa S., Fuchs H.R., Tarabykin V., Vida I, & Wulczyn F.G. (2015). miR-128 regulates neuronal migration, outgrowth and intrinsic excitability via the intellectual disability gene Phf6. eLife, 4, e04263.

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Immunohistochemical Analysis of Cortical Development

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Sections were dried at room temperature and incubated for 25 min in 10 mM sodium citrate buffer (pH 6.0) at 95°C. For BrdU staining, sections were treated with 2 N HCl for 30 min at room temperature. Sections were then blocked in 3% normal sheep serum and 0.1% Triton X-100 in PBS (blocking buffer) for 1 hour at room temperature. Then, sections were incubated in primary antibodies [rat anti-CTIP2 (1:500; Abcam, ab18465), mouse anti-SATB2 (1:500; Abcam, ab51502), rabbit anti-CUX1 (1:100; Santa Cruz Biotechnology, sc-13024), mouse anti-FOXP2 (1:250; Sigma-Aldrich, AMAB91361), rabbit anti-H3K36me3 (1:100; Abcam, ab9050), rabbit anti–cleaved Casp3 (1:100; Cell Signaling Technology, 9664S), rabbit anti-TBR2 (1:1000; Abcam, ab23345), and rat anti-BrdU (1:500; Abcam, ab6326)] in blocking buffer overnight at 4°C. After three times of rinsing in PBS, sections were incubated in secondary antibodies (Alexa Fluor 488–conjugated anti-mouse, Alexa Fluor 555–conjugated anti-mouse, Alexa Fluor 488–conjugated anti-rat, Alexa Fluor 488–conjugated anti-rabbit, and Alexa Fluor 555–conjugated anti-rabbit; Thermo Fisher Scientific; 1:1000) for 1 hour at room temperature. Nuclei were labeled by incubation in PBS containing 4′,6-diamidino-2-phenylindole (0.1 μg/ml) (Sigma-Aldrich), and samples were mounted in ProLong Gold Antifade Mountant (Thermo Fisher Scientific).

Xu L., Zheng Y., Li X., Wang A., Huo D., Li Q., Wang S., Luo Z., Liu Y., Xu F., Wu X., Wu M, & Zhou Y. (2021). Abnormal neocortex arealization and Sotos-like syndrome–associated behavior in Setd2 mutant mice. Science Advances, 7(1), eaba1180.

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Immunohistochemical Analysis of Murine Brain Tissue

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Brains were taken out immediately after perfusion and were postfixed for 1 h (P0), 1–2 h (P4–P14) or 1–4 h (P21). After the postfixation, the brains were washed in 0.1 m phosphate buffer for 20 min and were sunk in 30% sucrose/0.1 m phosphate buffer. Sections were cut as described above for in situ hybridization. The following primary antibodies were used: β-galactosidase (β-gal; 1:100, goat, catalog #55976, Cappel; 1:500, chicken, catalog #ab9361, Abcam); SOX6 (1:100, rabbit, catalog #ab30455, Abcam); SP8 (1:100, goat, catalog #sc-104661, Santa Cruz Biotechnology); CTIP2 (1:200, rat, catalog #ab18465, Abcam); TBR1 (1:200, rabbit, catalog #ab31940, Abcam; 1:200, chicken, catalog #AB2261, Millipore); PV (1:500, rabbit, catalog #PV27, SWANT); somatostatin (SST; 1:100, rat, catalog #MAB354, Millipore); LIM Homeobox 6 (LHX6; 1:50, mouse, catalog #sc-271433, Santa Cruz Biotechnology); vesicle-associated membrane protein 2 (VAMP2; 1:200, rabbit, catalog #104 202, Synaptic Systems); NetrinG1 (1:100, goat, catalog #AF1166, R&D Systems); and cleaved caspase 3 (1:100, rabbit, catalog #D175, Cell Signaling Technology). Secondary antibodies conjugated with Cy2, Cy3, or Cy5 were obtained from Jackson ImmunoResearch.

Larsen R., Proue A., Scott E.P., Christiansen M, & Nakagawa Y. (2019). The Thalamus Regulates Retinoic Acid Signaling and Development of Parvalbumin Interneurons in Postnatal Mouse Prefrontal Cortex. eNeuro, 6(1), ENEURO.0018-19.2019.

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Immunofluorescent Labeling of Neuronal Markers

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Cell cultures were fixed in 4% (w/v) paraformaldehyde in PBS and blocked with 5% normal donkey serum in 0.3% (v/v) Tween 20 in Tris-buffered saline before immunofluorescent staining. Primary antibodies used were anti-MAP2 (ab5392, Abcam), anti-CTIP2 (ab18465, Abcam), and anti-TBR1 (ab31940, Abcam), and secondary antibodies were Alexa Fluor conjugated. Stained cells were imaged on an Olympus FV1000 inverted confocal microscope and data imported into PerkinElmer Volocity for visualization.

Brownjohn P.W., Smith J., Portelius E., Serneels L., Kvartsberg H., De Strooper B., Blennow K., Zetterberg H, & Livesey F.J. (2017). Phenotypic Screening Identifies Modulators of Amyloid Precursor Protein Processing in Human Stem Cell Models of Alzheimer’s Disease. Stem Cell Reports, 8(4), 870-882.

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Immunofluorescent Labeling of Mouse Brain Sections

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Brains were harvested, postfixed in 4% paraformaldehyde, dehydrated with 30% sucrose in PBS, and serially sectioned at 40 μm on a vibratome (Leica). For immunofluorescence, sections were permeabilized in 0.1% Triton X-100/PBS for 10 min and blocked with 4% normal goat serum in PBS/0.1% Triton X-100 for 1 h at room temperature. Sections were then incubated with primary antibodies in 2% serum in PBS/0.1% Triton X-100 overnight at 4°C. The primary antibodies used in this study were rabbit anti-glial fibrillary acidic protein (GFAP; G9269, Sigma); chicken anti-neurofilament H (AB5539, Millipore Bioscience Research Reagents); rat anti-L1 (MAB5272, Millipore Bioscience Research Reagents); rat anti-Ctip2 (ab18465, AbCam), rabbit anti-APLP2 (gift from G. Thinakaran), APP Y188 (AbCam) and 6E10 (Covance). Sections were then washed and incubated with Alexa Fluor 488- or Alexa Fluor 596-conjugated secondary antibody (Invitrogen) for 1 h at room temperature. For myelin staining, mouse coronal sections were incubated with FluoroMyelin Green fluorescent myelin stain (1:300, Invitrogen) for 20 min at room temperature. After washing with PBS, sections were mounted with Vectashield Mounting Medium with DAPI (Vector Laboratories). Images were captured using a Leica confocal microscope.

Wang B., Li H., Mutlu S.A., Bowser D.A., Moore M.J., Wang M.C, & Zheng H. (2017). The Amyloid Precursor Protein Is a Conserved Receptor for Slit to Mediate Axon Guidance. eNeuro, 4(3), ENEURO.0185-17.2017.

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Immunohistochemical Labeling of Neural Markers

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Rat anti-BrdU (1:250, Abcam, #AB6326), rat anti-CTIP2 (1:500, Abcam, #AB18465), rabbit anti-CTIP2 (1:500, Abcam, #AB28448), rabbit anti-CUX1 (1:500, Santa Cruz, #sc-13024), rabbit anti-FITC (1:2000, Abcam, #AB19491), goat anti-FITC (1:1000, Novus Biolab, #NB600-493), mouse anti-SATB2 (1:200, Abcam, #AB51502), rabbit anti-TBR1 (1:500, Abcam, #AB31940).

Magrinelli E., Baumann N., Wagener R.J., Glangetas C., Bellone C., Jabaudon D, & Klingler E. (2022). Heterogeneous fates of simultaneously-born neurons in the cortical ventricular zone. Scientific Reports, 12, 6022.

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Quantitative Analysis of AAV-Transduced Neurons

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Based on the previous study (Lu and Yang, 2017 (link)), the AAV-infused mouse brains were sectioned at 60 μm of thickness. The sections were stained with GFP antibody (1:1000, ab6662; Abcam, Cambridge, UK) and Ctip2 antibody (1:500, ab18465, Abcam, Cambridge, UK). Afterward, the stained sections were imaged using a laser scanning confocal microscope (A1 confocal, Nikon Instruments [Shanghai]Co., Ltd.) under 40× objective lens. The MSNs were identified based on the positive staining of Ctip2. Neuronal structure reconstruction with neuTube (Feng et al., 2015 (link)) and Sholl analysis were performed with ImageJ (Longair et al., 2011 (link)).

Chen X., Yang Z., Shao Y., Kim K., Wang Y., Wang Y., Wu H., Xu X, & Le W. (2022). Pitx3 deficiency promotes age-dependent alterations in striatal medium spiny neurons. Frontiers in Aging Neuroscience, 14, 960479.

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Immunofluorescence Staining of Organoids and Brains

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Samples of organoids and mice brains were subjected to a standard pipeline for immunofluorescence staining, with the following primary antibodies: anti-PAX6 (Thermo Fisher, 42-6600, 1:400), anti-CTIP2 (Abcam, ab18465, 1:200), anti-SOX2 (R&D, AF2018, 1:400), anti-NEUN (Abcam; ab177484, 1:200), anti-TBR1 (Abcam, ab31940, 1:200), anti-CTIP2 (Abcam, ab18465, 1:200) and anti-SATB2 (Abcam, ab92446, 1:200). Images were taken under a confocal microscope (Carl Zeiss LSM880) and processed using ZEN 2012 (version 1.1.0.0). Cells were manually counted using ImageJ.

An N.A., Zhang J., Mo F., Luan X., Tian L., Shen Q.S., Li X., Li C., Zhou F., Zhang B., Ji M., Qi J., Zhou W.Z., Ding W., Chen J.Y., Yu J., Zhang L., Shu S., Hu B, & Li C.Y. (2023). De novo genes with an lncRNA origin encode unique human brain developmental functionality. Nature Ecology & Evolution, 7(2), 264-278.

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