Rabbit anti ng2

Manufactured by Abcam

Sourced in United Kingdom

Rabbit anti-NG2 is a primary antibody that recognizes the NG2 chondroitin sulfate proteoglycan, also known as CSPG4. NG2 is a transmembrane protein that is expressed on the surface of various cell types, including oligodendrocyte precursor cells, pericytes, and certain types of cancer cells. This antibody can be used to detect and study the expression and localization of NG2 in various biological samples.

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4 protocols using rabbit anti ng2

Immunofluorescent Staining of Neural Cells

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Cell cultures or brain slices were fixed in 4% paraformaldehyde in PBS, rinsed, permeabilized in 0.1% (for cells) or 3% (for slices) Triton X-100 in PBS, and then blocked with 5% normal goat serum (NGS) or normal donkey serum (NDS) in PBS containing 0.1% (for cells) or 0.3% (for slices) Triton X-100 for 1 h. Samples were then incubated with primary antibodies in blocking buffer, washed, and then probed with secondary antibodies (Alexa Fluor secondary antibodies, all from Life Technologies). After washing, the coverslips were mounted with ProLong Gold anti-fade reagent (Life Technologies). The brain slices were mounted with Fluoromount-G (Southern Biotech, Birmingham, AL), placed under the glass covers, and sealed. Primary antibodies used in the immunostaining were mouse anti-NeuN (Millipore), mouse anti-β-tubulin (Sigma), rabbit anti-GFAP (DAKO, Carpinteria, CA), mouse anti-Olig1(Millipore), mouse anti-O4 IgM (R&D Systems, Minneapolis, MN ), rabbit anti-LAMP1(Abcam, Cambridge, UK), rabbit anti-Calbindin (Millipore), mouse anti-C3d (a gift from Dr. Joshua Thurman, University of Colorado), rabbit anti-MAC complex (Abcam), and rabbit anti-NG2 and rabbit anti-Olig2 (both are gifts from Dr. Charles Stiles, Harvard University).

Liu Y., Harlow D.E., Given K.S., Owens G.P., Macklin W.B, & Bennett J.L. (2016). Variable sensitivity to complement-dependent cytotoxicity in murine models of neuromyelitis optica. Journal of Neuroinflammation, 13, 301.

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Western Blot Analysis of Neural Markers

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The concentration of protein was determined by BCA method to prepare protein loading samples. The brain protein extract (30 μg) was separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and subsequently transferred to the PVDF membrane. The membrane was blocked with 5% skim milk on a shaker for 1 h. After washing with TBST, the membranes were incubated with the primary antibodies rabbit anti-MBP (1:500, Abcam), goat anti-Iba-1 (1:500, Abcam), rabbit anti-Arg-1 (1:800, Cell Signaling Technology), rabbit anti-iNOS(1:500, Genetex), rabbit anti-BDNF (1:500, Abclonal), rabbit anti-GDNF(1:500, Abcam), rabbit anti-CNTF (1:1000, Abcam), rabbit anti-NG2 (1:1000, Abcam), rabbit anti-β-actin (1:1000, Abclonal) and rabbit anti-Tubulin (1:1000, Abclonal) overnight at 4 °C. After washing with TBST, the membranes were added with the HRP–conjugated goat anti-rabbit (1:3000, Abclonal) and rabbit anti-goat (1:1000, Boster) second antibodies for 2 h at RT. After washing with TBST, immunoblots were developed with an enhanced chemiluminescence systemand measured using Quantity Software (Bio-Rad, Hercules, CA, USA).

Li Y.R., Sun M.Y., Hang W., Xiao Q., Fan H.J., Jia L., Jin X.M., Zhang B., Xiao B.G., Ma C.G, & Chai Z. (2022). Wuzi Yanzong Pill relieves CPZ-induced demyelination by improving the microenvironment in the brain. Heliyon, 8(12), e12277.

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Immunofluorescence Labeling of SCN Slices

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Antiserum for immunofluorescence on SCN slices: goat anti-glutamate transporter neuronal EAAT3 Millipore 1:1000; rabbit anti-GAD65/67 1:1000 Abcam; rabbit anti-AVP, Bachem 1:500; rabbit anti-Aldh1L1 1:200 Abcam; rabbit anti-NG2 1:200 Abcam; goat anti-Iba1 1:350 Wako; Secondary antibodies: donkey anti-goat conjugated with Alexa 647; donkey anti-goat conjugated with Alexa 488; chicken anti-rabbit conjugated with Alexa 488; goat anti-rabbit conjugated with Alexa 488; goat anti-rabbit conjugated with Alexa 647 (Life Technologies). Targeting rates and co-localization analysis of Syn-mCherry::Cre and GFAP-EGFP::Cre co-transduced SCN slice was manually performed on single confocal planes. Number of DAPI⁺ cells was also assessed manually, because of natural unevenness of DAPI staining in thick slices. Co-localization of GFAP-mCherry::Cre with the different astroglial markers (Aldh1L1, Iba1, NG2) was also manually performed on single confocal planes.

Brancaccio M., Patton A.P., Chesham J.E., Maywood E.S, & Hastings M.H. (2017). Astrocytes Control Circadian Timekeeping in the Suprachiasmatic Nucleus via Glutamatergic Signaling. Neuron, 93(6), 1420-1435.e5.

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Profiling Corpus Callosum Protein Markers

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The corpus callosum of mice was carefully separated and then lysed with homogenization buffer. The cells were homogenized in RIPA buffer. Equal amounts (40 µg) of tissue lysates were separated using SDS-PAGE and transferred onto a nitrocellulose membrane. The membranes were incubated with the following primary antibodies: rabbit anti-H₂R (1:500, Abcam); rat anti-MBP (1:1,000, Millipore); rabbit anti-NG2 (1:1,000, Abcam); mouse anti-CC-1 (1:1,000, Abcam); rabbit β-catenin (1:300, Abcam); rabbit GSK3β (1:1,000, Abcam); rabbit GSK3β (phosphor Y216; 1:1,000, Abcam); rabbit cleaved-caspase3 (1:1,000, Abcam); rabbit CREB (1:500, Abcam); rabbit p-CREB (1:500, Abcam) and mouse anti-GAPDH (1:5,000, Kang-chen). Secondary antibodies conjugated with HRP against rabbit, mouse, or rat IgG (1:3,000, Lianke) were applied. Images were acquired with the Tanon Chemiluminescent Imaging System (Tanon) and analyzed by ImageJ software (National Institutes of Health). The results were expressed as the target protein/GAPDH ratio and then normalized to the values measured in the sham or control groups in vivo and in vitro.

Jiang L., Cheng L., Chen H., Dai H., An D., Ma Q., Zheng Y., Zhang X., Hu W, & Chen Z. (2020). Histamine H2 receptor negatively regulates oligodendrocyte differentiation in neonatal hypoxic-ischemic white matter injury. The Journal of Experimental Medicine, 218(1), e20191365.

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