Mouse anti o4

Manufactured by R&D Systems

Sourced in United States

Mouse anti-O4 is a monoclonal antibody that recognizes the O4 antigen, which is expressed on the surface of oligodendrocyte precursor cells and immature oligodendrocytes. This antibody can be used for the identification and isolation of these cell types.

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Lab products found in correlation

Rat anti gfap, by Thermo Fisher Scientific (2 mentions) Goat anti iba 1, by Novus Biologicals (2 mentions) Rabbit anti aldolase c, by Thermo Fisher Scientific (2 mentions) Rabbit anti gfap, by Agilent Technologies (2 mentions) Mouse anti o1, by R&D Systems (2 mentions) Mouse anti tuj1, by Merck Group (2 mentions) Anti mouse alexa fluor 488, by Thermo Fisher Scientific (2 mentions) Rabbit anti gfap, by Merck Group (1 mentions) Mouse anti ed1, by Bio-Rad (1 mentions) Mouse anti reca 1, by Abcam (1 mentions) Abc elite kit, by Vector Laboratories (1 mentions) Ix70 microscope, by Olympus (1 mentions) Eclipse te 2000 e2, by Nikon (1 mentions) Goat anti nestin, by Santa Cruz Biotechnology (1 mentions) Rabbit anti sox2, by Abcam (1 mentions) Rabbit anti tbr2, by Merck Group (1 mentions) Mouse anti neun, by Merck Group (1 mentions) Photomicroscope, by Olympus (1 mentions) Alexa flour conjugated secondary antibody, by Thermo Fisher Scientific (1 mentions) Goat serum, by Thermo Fisher Scientific (1 mentions)

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Anti-O4 (5 citations)

7 protocols using mouse anti o4

Cytospin-Based Immunocytochemistry of Sorted tdT+ Cells

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For preparation mounting FACS-sorted tdT + cells Cytospin centrifugation was used. TdT positive cells were suspended in 100 μL of neurosphere media containing DMEM/F-12 supplemented with 5 μg/mL Heparin, N2, and B27. Those cells were placed on charged glass histology slide with filter adapter for the cytospin centrifuge. After spinning at 300 rpm for 5 min, sides were airdried for 15 min followed by fixation with 4% PFA at room temperature for 15 min. Immunocytochemistry was then performed as described below.
Immunocytochemistry was performed according to the following protocol. After fixation cells were incubated in blocking buffer (5% donkey serum and 0.2% Triton X-100) for 45 min. Cells were then incubated in primary antibodies diluted in blocking buffer at 4°C overnight. Appropriate secondary antibodies were used for single and double labeling. All secondary antibodies were tested for cross-reactivity and nonspecific immunoreactivity. The following primary antibodies were used: mouse anti-beta III-tubulin (TUJ1), (Millipore; cat #MAB1637; 1:500), chicken anti-MAP2 (Millipore; cat #AB5543; 1:500), rat anti-GFAP (Invitrogen; cat #13–0300; 1:400), rabbit anti-Aldolase C (Thermofisher; cat #14884-1-AP; 1:100), mouse anti-O4 (R&D; cat #MAB1326; 1:400) and goat anti-IBA-1(Novus; cat #nb100-1028; 1:400).

Wei H., Wu X., Withrow J., Duran R.C., Singh S., Chaboub L.S., Rakshit J., Mejia J., Rolfe A., Herrera J.J., Horner P.J, & Wu J.Q. (2023). Glial progenitor heterogeneity and key regulators revealed by single-cell RNA sequencing provide insight to regeneration in spinal cord injury. Cell reports, 42(5), 112486.

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Immunocytochemical Staining of Neural Cells

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Cultures were fixed in 4 % paraformaldehyde, as previously described [18 (link)]. Background staining was reduced by blocking nonspecific binding sites with 10 % goat serum for 1 h at room temperature. After washing with phosphate-buffered saline (PBS), endogenous peroxidase activity was quenched by incubation with a 3 % H₂O₂ solution in PBS. Cultures were washed again and then incubated overnight with the appropriate primary antibodies (rabbit anti-MAP-2, 1:500, Chemicon; rabbit anti-GFAP, 1:1000, Chemicon; mouse anti-ED1, 1:500, Serotec; mouse anti-O4, 1:300, R&D system; rabbit-anti fibronectin 1, 1:500, Bioworld Technology; mouse-anti RECA-1, 1:300, Abcam) at 4 °C. Cells were washed and visualized using the avidin-biotin peroxidase complex method (ABC Elite kit; Vector Laboratories, Burlingame, CA). Images were viewed on an inverted Olympus IX 70 microscope (Tokyo City, Japan) equipped with a cooled CCD camera and SPOT advanced software (Diagnostic Instruments, Sterling Heights, MI).

Huang Y.N., Ho Y.J., Lai C.C., Chiu C.T, & Wang J.Y. (2015). 1,25-Dihydroxyvitamin D3 attenuates endotoxin-induced production of inflammatory mediators by inhibiting MAPK activation in primary cortical neuron-glia cultures. Journal of Neuroinflammation, 12, 147.

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Immunohistochemical Analysis of Neural Cell Markers

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These were performed as described previously [37 (link)]. Frozen coronal brain sections (10–14 μm) were prepared with a cryostat for immunohistochemistry. The following primary antibodies were used: mouse anti-bromodeoxyuridine (BrdU) (DAKO Cytomation), rabbit anti-GFAP (DAKO), goat anti-Nestin (Santa Cruz Biotech, Dallas, TX, www.scbt.com), mouse anti-NeuN (Millipore, Billerica, MA, www.millipore.com), mouse anti-O4 (R&D, Minneapolis, MN, www.rndsystems.com), rabbit anti-Sox2 (Epitomics, Burlingame, CA, www.epitomics.com), rabbit anti-Tbr2 (Millipore), and mouse anti-Tuj1 (Millipore). All images were collected by a laser scanning spectral confocal microscope system (Nikon Eclipse TE2000-E2, Tokyo, Japan, www.nikon.com) and processed with the Adobe Photoshop 7.0 software (Adobe Systems, San Jose, CA, www.adobe.com).

Gan Q., Lee A., Suzuki R., Yamagami T., Stokes A., Nguyen B.C., Pleasure D., Wang J., Chen H, & Zhou C.J. (2014). Pax6 Mediates β-Catenin Signaling for Self-Renewal and Neurogenesis by Neocortical Radial Glial Stem Cells. Stem cells (Dayton, Ohio), 32(1), 45-58.

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Immunohistochemistry of Cultured Cells and Brain Slices

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Cultured cells or brain slices (30 μm) were fixed with 4% paraformaldehyde followed by three washes in PBS and permeabilized with 0.1% Triton X-100 for 10 min (except for O4, O1 immunostainning) then blocked in 10% goat serum (Invitrogen Corp. USA) for 1 h following incubation with the primary antibody overnight at 4 °C. Primary antibodies were diluted in blocking solutions as follows: mouse anti-MBP (Biolegend, USA; 1:500), mouse anti-APC, CC1 clone (Millipore, USA; 1:500), mouse anti-O4 (R&D Systems Inc, USA, 1:800), mouse anti-O1 (R&D Systems Inc, USA, 1:800), rabbit anti-PDGFRα (Cell Signalling Technology, USA; 1:500), rabbit anti-glial fibrillary acidic protein (GFAP, Biolegend, USA; 1:500) mouse anti-BrdU (Sigma St Louis, USA; 1:500). The tissue or cells were then washed with PBS and incubate in Alexa Flour-conjugated secondary antibodies (Invitrogen Corp. USA; 1:500) for 1 h. Images were obtained using Olympus photomicroscope and analyzed using Image-Pro plus 6.0 software.

Yu X., Cheng G., Zhang L., Zhang Y., Wang Q., Zhao M., Zeng L., Hu Y, & Feng L. (2018). N-Phenylquinazolin-2-amine Yhhu4952 as a novel promotor for oligodendrocyte differentiation and myelination. Scientific Reports, 8, 14040.

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Immunocytochemistry of Oligodendrocyte Markers

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The following primary antibodies were used for immunocytochemistry at the indicated dilutions: mouse anti‐O1 (1 : 500; R&D systems, Minneapolis, MN, USA, cat# MAB1327, RRID:AB_357618), mouse anti‐O4 (1 : 500; R&D systems, cat# MAB1326, RRID:AB_357617), and rabbit anti‐SLC7A11 (1 : 100; ProteinTech, Chicago, IL, USA, cat# 26864‐1‐AP). The secondary antibodies used were as follows: anti‐mouse Alexa Fluor 488 (1 : 1000; Thermo Fisher Scientific, Waltham, MA, USA, cat# A21202, RRID: AB_141607) and anti‐rabbit Alexa Fluor 647 (1 : 1000; Thermo Fisher Scientific, cat# A31573, AB_2536183).

Hoshino T., Yamakado H., Takahashi R, & Matsuzawa S. (2020). Susceptibility to erastin‐induced ferroptosis decreases during maturation in a human oligodendrocyte cell line. FEBS Open Bio, 10(9), 1758-1764.

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Cytospin-Based Immunocytochemistry of Sorted tdT+ Cells

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Detailed Immunohistochemical Labeling Protocol

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All animals were deeply anesthetized, perfused, and stained as previously described [7 (link)]. After perfusion, the brain was isolated, post-fixed overnight with 4% PFA, and sectioned with a vibratome at 50 μm intervals by embedding in 3% agarose in PBS (Leica, VT1000S). All sections through the hippocampus were collected in PBS containing 0.1% NaN₃ and stored at 4°C. Every twelfth section of brain tissue was permeabilized with 0.3% Triton X-100 in PBS, blocked for at least 2 hours at room temperature with 10% normal goat serum, and incubated with primary antibodies followed by secondary antibodies. For immunocytochemistry analysis, cells were fixed with 4% PFA for 20 minutes at room temperature, blocked with 10% normal goat serum in PBS containing 0.1% Tween20 for 1 hour, and incubated with primary then secondary antibodies. The primary antibodies used were: rabbit anti-GFAP (DAKO, 1:2000), rabbit anti-GFP (Proteintech, 1:500), rat anti-BrdU (Abcam, 1:500), mouse anti-PCNA (Santa Cruz, 1:200), mouse anti-Tuj1 (Sigma, 1:1000), rabbit anti-MAP2 (Millipore, 1:1000), rabbit anti-GFAP (DAKO, 1:1000), and mouse anti-O4 (R&D System, 1:1000). Fluorescent-conjugated secondary antibodies were used (Invitrogen, 1:1000). Biotinylated-conjugated anti-species IgG (Vector Laboratories, 1:500) were used for peroxidase/diaminobenzidine (DAB) staining in stereology analysis.

Liu B., Cheng W., Cheng D., Pu J., Nie Z., Xia C., Chen Y, & Yang C. (2021). PirB functions as an intrinsic suppressor in hippocampal neural stem cells. Aging (Albany NY), 13(12), 16062-16071.

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