Mouse anti mbp

Manufactured by Fortrea

Sourced in United Kingdom

Mouse anti-MBP is a laboratory reagent used in research applications. It is an antibody that specifically binds to the myelin basic protein (MBP) found in the myelin sheath of nerve cells. This antibody can be used for the detection and analysis of MBP in various experimental techniques.

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Lab products found in correlation

Rabbit anti olig2, by Abcam (3 mentions) Mouse anti plp, by Abcam (2 mentions) Rabbit anti gfap, by Agilent Technologies (2 mentions) Rabbit anti iba1, by Fujifilm (2 mentions) Rabbit anti olig2, by Merck Group (2 mentions) μ columns, by Miltenyi Biotec (2 mentions) Rat anti mbp, by Bio-Rad (2 mentions) Protein g microbeads, by Miltenyi Biotec (2 mentions) Mouse anti actin, by Merck Group (2 mentions) Donkey anti mouse alexa 488, by Thermo Fisher Scientific (1 mentions) Colibri light source, by Zeiss (1 mentions) Mowiol, by Merck Group (1 mentions) Zen 2.6 blue software, by Zeiss (1 mentions) Mrm camera, by Zeiss (1 mentions) Illustrator 2020, by Adobe (1 mentions) 4 6 diamidino 2 phenylindole (dapi), by Merck Group (1 mentions) Rabbit anti tuj1, by Fortrea (1 mentions) Kwikquant, by Kindle Biosciences (1 mentions) Mouse anti mog, by Merck Group (1 mentions) Smi 99p 100, by Merck Group (1 mentions)

13 protocols using mouse anti mbp

Immunostaining of Myelination and Neurite Outgrowth

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Cells from SC-DRG co-cultures were fixed in 4% paraformaldehyde (PFA) in PBS for 10 min and then permeabilized in a mixture of 95% ice-cold methanol and 5% acetone at −20 °C for 5 min. Afterwards cells were incubated in blocking solution (2% horse serum, 2% bovine serum albumin (BSA), 0.1% porcine gelantine) for 1 h at room temperature before incubation in primary antibodies (mouse anti-MBP 1:500 (Covance), rabbit anti-TUJ1 1:250 (Covance)) overnight at 4 °C. The next day, cells were washed in PBS for three times and incubated in secondary antibodies (Alexa 488 donkey anti mouse 1:1000 (Invitrogen) and Alexa 568 donkey anti rabbit 1:1000 (Invitrogen) diluted in blocking solution with 0.2 µg/µl 4’,6’-diamidino-2-phenylindole (DAPI; Sigma) for 1 h at room temperature. Following three washing steps in PBS cells were mounted on slides in Mowiol mounting solution (9.6% Mowiol (Sigma), 24% Glycerol, 0.1 M Tris-HCl). Fluorescent images were taken using a Axiophot Observer Z (Zeiss) with a Colibri light source (Zeiss) and MRM camera (Zeiss). Acquisition and processing of the images was carried out using Zen2.6 blue software (Zeiss), FIJI (NIH) and Illustrator 2020 (Adobe).

Krauter D., Stausberg D., Hartmann T.J., Volkmann S., Kungl T., Rasche D.A., Saher G., Fledrich R., Stassart R.M., Nave K.A., Goebbels S., Ewers D, & Sereda M.W. (2024). Targeting PI3K/Akt/mTOR signaling in rodent models of PMP22 gene-dosage diseases. EMBO Molecular Medicine, 16(3), 616-640.

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Multimodal Analysis of Spinal Cord Cell Types

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Spinal cords were fixed in 4% paraformaldehyde, and then cryoprotected in 20% sucrose. Mouse spinal cord was analyzed using colorimetric in situ hybridization as previously described (Kang et al., 2012 (link)). Immunohistochemistry: mouse anti-MBP (Covance 1:500), mouse anti-PLP (1:500), rabbit anti-Olig2 (Abcam 1:1000), rabbit anti-GFAP (DAKO 1:1000), rabbit anti-Iba1 (Wako 1:600).
To perform the two-color fluorescent in situ hybridization using the TSA-FITC/TSA-Cy5 system, we generated FITC labeled Apcdd1 probes, and combined with the probes of DIG labeled oligodendrocyte lineage markers (PDGFRα, PLP). Day 1 of the two-color fluorescent in situ hybridization was performed the same as for the colorimetric in situ. For day 2 of in situ, the slides were washed with 0.1X SSC and endogenous peroxidase activity was quenched by incubation in 3% H₂O₂ for 20 minutes. After quenching step, the slides were blocked with 0.5% of a blocking reagent containing TN (100 mM Tris-HCl, pH 7.5, 150 mM NaCl). The following antibodies were used to detect either FITC or DIG labeled probes; anti-FITC-POD, along with FITC-Tyramide; anti-DIG-POD with Cy5-Tyramide in Amplification Reagent (TSA^™ kit).

Lee H.K., Laug D., Zhu W., Patel J.M., Ung K., Arenkiel B.R., Fancy S.P., Mohila C, & Deneen B. (2015). Apcdd1 stimulates oligodendrocyte differentiation after white matter injury. Glia, 63(10), 1840-1849.

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Western Blotting Characterization of Oligodendrocyte Markers

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Tissue samples were prepared, run on SDS-PAGE gels and transferred to PVDF membranes as described^{34 (link)}. Transferred membranes were incubated with primary antibodies overnight at 4°C. Primary antibodies used were as follows: Rabbit anti-Olig2 (1:2,000 Millipore AB9610), Mouse anti-MBP (1:2,000 Covance SMI-99P-100), Mouse anti-MOG (1:3,000 Millipore MAB5680), Rabbit anti-Neurofilament H (1:10,000 Millipore AB1989), Rabbit anti-GDE2 (1:1000), Rabbit anti-PDGF receptor α (1:2,000 Cell Signaling Technology 3174), Mouse anti-Actin (1:10,000 Millipore MAB1501). After 1 hour incubation at room temperature with appropriate HRP-conjugated secondary antibodies, membranes were developed by film or by using a digital imaging system (KwikQuant, Kindle Biosciences).

Choi B.R., Dobrowolski M, & Sockanathan S. (2020). GDE2 expression in oligodendroglia regulates the pace of oligodendrocyte maturation. Developmental dynamics : an official publication of the American Association of Anatomists, 250(4), 513-526.

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Quantitative Protein Analysis via Western Blot

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Gel electrophoresis and Western blotting were performed as described in Kosturko et al., (2005) (link) using the following antibodies: mouse anti-MBP (Covance, 1:10,000 dilution), mouse anti-myelin oligodendrocyte glycoprotein (MOG) (Millipore, 1:1000 dilution), and mouse anti-proteolipid protein (PLP) (Millipore 1:1,000 dilution), and appropriate horseradish conjugated secondary antibodies (Jackson ImmunoResearch, 1:10,000 dilution). Western blots were visualized using chemiluminescence detection (Pierce). Densitometric analysis was performed using Adobe Photoshop. Optical intensity of bands was integrated after subtracting background; levels of test proteins were normalized to β-actin or nucleoporin protein levels. The levels of proteins from control mice were set at 100%.

Maggipinto M.J., Ford J., Le K.H., Tutolo J.W., Furusho M., Wizeman J.W., Bansal R, & Barbarese E. (2017). Conditional knockout of TOG results in CNS hypomyelination. Glia, 65(3), 489-501.

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Immunofluorescence Analysis of Oligodendrocytes

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Tissue sections and cultured cells were fixed with 4% paraformaldehyde, permeabilized with detergent, incubated successively with blocking reagent, and with primary and secondary antibodies. The following primary antibodies were used: mouse anti-CNP (EMD Millipore, 1:200 dilution), mouse anti-MBP (Covance, 1:2,000 dilution), rabbit anti-OLIG2 (Abcam, 1:1000 dilution) and rabbit anti-TOG antibody (Abcam, 1:400 dilution). Secondary conjugated antibodies were from Molecular Probes. Nuclei were stained with TO-PRO-3 (1:500) (Invitrogen). Tissue sections and cultured cells were covered with Prolong Gold antifade reagent (Invitrogen). Fluorescent images were acquired by confocal microscopy.

Maggipinto M.J., Ford J., Le K.H., Tutolo J.W., Furusho M., Wizeman J.W., Bansal R, & Barbarese E. (2017). Conditional knockout of TOG results in CNS hypomyelination. Glia, 65(3), 489-501.

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Immunofluorescence and Western Blot Analyses of Erk1/2, MBP, and Olig2 Markers

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The following primary antibodies were used for immunofluorescence analyses: mouse anti-pErk1/2 (1:300, Cell Signaling #9106), mouse anti-MBP (1:300, Covance #SMI-94R), rabbit anti-Olig2 (1:10,000, kind gift from Dr. Charles Stiles, Dana-Farber Cancer Institute), and chicken anti-tubulin (1:1000, Aves #TUJ). Secondary antibodies were donkey anti-mouse Alexa 488 (Jackson ImmunoResearch #715-545-151); donkey anti-mouse rhodamine-X (Jackson ImmunoResearch #715-295-151); donkey antichicken Alex 488 (Jackson ImmunoResearch #703-545-155). All secondary antibodies were used at 1:250. The following primary antibodies were used for Western blot analyses: mouse anti-pErk1/2 (1:1000, Cell Signaling #5726), rabbit anti-total Erk1/2 (1:1000, Promega #V114A), rabbit anti-MBP (1:1000, Chemicon #AB980) and mouse anti-ß-actin (1:5000, Sigma #A5441).

Kim J., Adams A.A., Gokina P., Zambrano B., Jayakumaran J., Dobrowolski R., Maurel P., Pfister B.J, & Kim H.A. (2020). Mechanical stretch induces myelin protein loss in oligodendrocytes by activating Erk1/2 in a calcium-dependent manner. Glia, 68(10), 2070-2085.

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Immunohistochemistry of Neural Cell Markers

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Antibodies used in this study were: rabbit anti-NG2 (1:400, Millipore), mouse anti-CC1 (CC-1, 1:400, MERC), sheep anti-BrdU (1:400, Exalpha Biologicals), mouse anti-MBP (1:4000, Covance), goat anti-Notch1ICD (1:50, Santa Cruz), goat anti-Prox1 (1:50, R & D system), rat anti-PDGFRα (1:100, eBioscience), rat F4/80 (1:1000, Serotec), chick anti-GFP (1:2000, Aves), mouse anti-GFP (1:400, Life Technologies), rabbit anti-GFP (1:500, Life Technologies), Alexa 488, 594, 647 conjugated donkey secondary antibodies (1:400, Life Technologies), biotinylated donkey anti-goat and biotinylated donkey anti-chicken (1:400, Life Technologies), and Streptavidin 488, 546 (1:400, Life Technologies).

Kato K., Konno D., Berry M., Matsuzaki F., Logan A, & Hidalgo A. (2015). Prox1 Inhibits Proliferation and Is Required for Differentiation of the Oligodendrocyte Cell Lineage in the Mouse. PLoS ONE, 10(12), e0145334.

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Isolating and Differentiating Oligodendrocyte Progenitor Cells

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OPCs were gently removed from the fibronectin-coated plates using solution containing 44 U/mL papain (Worthington) and DNase I in MEM with 20 mM Hepes, 5 mM EDTA and 14 mM L-cysteine. The solution was swirled over the cells and then aspirated to leave a thin film of solution. The cells were returned to the incubator for 7 min. Plates were agitated by hitting vigorously. Once the cells were detached, 5 mL of N2B2 was added and the cells were collected by centrifugation at 400 x g for 8 min. The pellet was resuspended and viable cells were plated in N2S on fibronectincoated flasks at no less than 2.5x10 4 cells/cm 2 . Again, half media changes were performed every other day until the cells were 80% confluent and ready to be passaged. To differentiate the OPCs, the mitogens were removed from the media and the cells were placed into N2B2 supplemented with 1.95 μg/mL triiodothyronine (T3) (Sigma) [27, 28] , 1 ng/mL recombinant rat CNTF (R&D Systems) [29] , and 1 ng/mL TGFß1 (R&D Systems) [30] for 5 -7 days[LMM1]. Then stained for mouse anti-O4 (1:4, supernatant), mouse anti-O1 (1:4, supernatant), rat anti-PLP (1:1000, gift from Wendy Macklin), and mouse anti-MBP (1:250, Covance), rabbit anti-Olig2 (1:100, Milipore), and rat anti-GF2.2 (1:4, supernatant).

Moore L., McLane L.E., Wahl S., Ornelas I.M., Wood T.L., Canoll P., & Levison S.W. (2021). Hyperoxia Inhibits the Growth of Mouse Forebrain Oligodendrocyte Progenitors.

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Cerebellar Slice Immunohistochemistry Protocol

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After treatment, cerebellar slices were rinsed twice with PBS and fixed in 4% paraformaldehyde in PBS for 20 min at room temperature. For immunohistochemistry, slices were rinsed in PBS and permeabilized in 1.5% or 10% (myelin proteins) Triton X-100 in PBS for 20 min. Slices were rinsed, blocked with 5% normal donkey serum (NDS) in PBS with 0.3% Triton X-100 for 1 h, and incubated with primary antibodies overnight at room temperature. Following 3 washes in PBS, Alexa Fluor-labeled secondary antibodies (Jackson ImmunoResearch, West Grove, PA) were applied (1:800) overnight at room temperature, washed 3 times in PBS and mounted in Fluoromount G (Southern Biotech, Birmingham, AL). The following primary antibodies were used: goat anti-Sox10 (Santa Cruz), rat anti-PLP (Yamamura, Konola, Wekerle, & Lees, 1991 (link)), mouse anti-AQP4 (Santa Cruz Biotechnology, Dallas, TX), rabbit anti-S100b (Sigma), rabbit anti-GST-pi (Enzo), mouse anti-Caspr (NeuroMAB), mouse anti-CC1 (Calbiochem), rabbit anti-GFAP (Sigma), rabbit anti-MOG (Abcam, Cambridge, United Kingdom), mouse anti-MBP (Covance, Princeton, NJ), chicken anti- Neurofilament-H (Neuromics, Minneapolis, MN), goat anti-Iba1 (Abcam).

Liu Y., Given K.S., Owens G.P., Macklin W.B, & Bennett J.L. (2018). Distinct Patterns of Glia Repair and Remyelination in Antibody-mediated Demyelination Models of Multiple Sclerosis and Neuromyelitis Optica. Glia, 66(12), 2575-2588.

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Spinal Cord Immunohistochemistry for Cell Markers

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Spinal cords were dissected from the spinal columns and paraffin sections from the lumbar region were prepared and analyzed for changes in the expression of CD45, CD3, and myelin basic protein (MBP) using multi-channel fluorescence immunohistochemistry using immunostaining methods as previously described.^{50 (link)} Rabbit-anti CD45 (1:50, Abcam), rat anti-CD3 (1:100, Abcam) and mouse anti-MBP (1:1000; Covance) and appropriate fluoroconjugated secondary antibodies (Alexa 546 or Alexa 488, Molecular Probes, Inc.) were used. Cell nuclei were visualized by staining with DAPI (1:5000, Molecular Probes, Inc.). Sections were photographed with an Axioskop 40 fluorescence microscope (Zeiss) and images processed under identical conditions using Image J (version 1.53t).

Verbout N.G., Su W., Pham P., Jordan K., Kohs T.C., Tucker E.I., McCarty O.J, & Sherman L.S. (2023). E-WE thrombin, a protein C activator, reduces disease severity and spinal cord inflammation in relapsing-remitting murine experimental autoimmune encephalomyelitis. Research Square.

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