Nf200 antibody

Manufactured by Merck Group

Sourced in Israel, United States, Japan

The NF200 antibody is a laboratory reagent used for the detection and analysis of neurofilament proteins. It is a widely used tool in neuroscience research and neurological disease studies. The NF200 antibody specifically recognizes the neurofilament heavy chain, which is a key structural component of neurons. This product can be used in various experimental techniques, such as immunohistochemistry, Western blotting, and flow cytometry, to identify and quantify neurofilament proteins in biological samples.

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Lab products found in correlation

Arsenite, by Merck Group (2 mentions) Goat serum, by Vector Laboratories (2 mentions) Fetal calf serum (fcs), by Thermo Fisher Scientific (2 mentions) Dulbecco s modified eagle s medium dmem, by Thermo Fisher Scientific (2 mentions) Dapi staining, by Vector Laboratories (2 mentions) Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions) Glutamax, by Thermo Fisher Scientific (2 mentions) Paraformaldehyde (pfa), by Merck Group (2 mentions) Triton x 100, by Merck Group (2 mentions) Phosphate buffered saline (pbs), by Merck Group (2 mentions) Fugene 6, by Promega (2 mentions) Alexa fluor 488 donkey anti rabbit igg, by Thermo Fisher Scientific (1 mentions) Alexa fluor 594 goat anti mouse igg, by Thermo Fisher Scientific (1 mentions) Cleaved caspase 3 antibody, by Cell Signaling Technology (1 mentions) Alexa fluor 488 goat anti mouse igg, by Proteintech (1 mentions) Immunol staining blocking buffer, by Beyotime (1 mentions) Sa00009 2, by Proteintech (1 mentions) Axio imager m2, by Zeiss (1 mentions) Shandon m1 embedding matrix, by Thermo Fisher Scientific (1 mentions) Sodium azide, by Merck Group (1 mentions)

8 protocols using nf200 antibody

Immunofluorescence Staining of Dorsal Root Ganglia

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Rats were perfused with 4% paraformaldehyde, and DRG was extracted and postfixed
overnight in 4% paraformaldehyde. Immunofluorescence staining was performed on
cryosections (16 µm) using standard reagents and protocols. Briefly, the sections were
incubated with a solution containing 0.3% Triton X-100 and 1% bovine serum albumin for
30 min at room temperature. The sections then were incubated with NF200 antibody (1:200,
Sigma) and anti-HCN1, anti-HCN2, anti-HCN3 antibody, respectively (1:200, Alomone, Israel)
for 24 h. After three rinses, the sections were further incubated with the secondary
antibodies Alexa Fluor 594 (goat anti-mouse IgG, 1:500, Invitrogen) and Alexa Fluor 488
(donkey anti-rabbit IgG, 1:500, Invitrogen) for 4 h at room temperature. The sections were
then mounted with anti-fade medium and stored at 4℃. All images were captured with an
Olympus confocal microscope and processed with Adobe Photoshop software.

Liu D.L., Wang X., Chu W.G., Lu N., Han W.J., Du Y.K., Hu S.J., Bai Z.T., Wu S.X., Xie R.G, & Luo C. (2017). Chronic cervical radiculopathic pain is associated with increased excitability and hyperpolarization-activated current (Ih) in large-diameter dorsal root ganglion neurons. Molecular Pain, 13, 1744806917707127.

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Immunohistochemical Staining Protocols

Cited 3 times

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The staining methods were previously described (Zhu et al., 2007 (link), 2009 (link), 2013 (link); Peng et al., 2012 (link)). The antibodies for staining were purchased from the following sources: IL-17c antibody (mouse monoclonal, R&D Systems); IL-17RE antibody (rabbit polyclonal, Sigma-Aldrich); IL-17RA antibody (rabbit monoclonal, LifeSpanBioSciences); NCAM antibody (mouse monoclonal, BD); PGP9.5 (Abcam); NF200 antibody (rabbit polyclonal, Sigma-Aldrich); Peripherin antibody (rabbit and mouse, Sigma-Aldrich); cleaved caspase 3 antibody (rabbit polyclonal, Cell Signaling Technology).

Peng T., Chanthaphavong R.S., Sun S., Trigilio J.A., Phasouk K., Jin L., Layton E.D., Li A.Z., Correnti C.E., De van der Schueren W., Vazquez J., O’Day D.R., Glass I.A., Knipe D.M., Wald A., Corey L, & Zhu J. (2017). Keratinocytes produce IL-17c to protect peripheral nervous systems during human HSV-2 reactivation. The Journal of Experimental Medicine, 214(8), 2315-2329.

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Immunofluorescence Analysis of Myelination

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Sciatic nerve tissues were fixed with 4% paraformaldehyde, washed with PBS, and blocked with Immunol Staining Blocking Buffer (Beyotime), incubated with primary MBP antibody (Cell Signaling Technology, 78896S, Boston, MA, USA), S100β antibody (Sigma, S2532) and NF200 antibody (Sigma, N2912), and then incubated with Goat Anti-Rabbit secondary antibody conjugated to Cy3 (Proteintech, SA00009-2, Chicago, IL, USA) and Alexa Fluor 488 Goat Anti-Mouse IgG (Proteintech, SA00013-1) prior to mounting on slides. Fluorescent signals from different lasers were used to visualize tissue slides. Optical and epifluorescence microscopes (Axio Imager M2, Carl Zeiss Microscopy GmbH, Jena, Germany) were used to capture the images.

Li S., Wu W., Zhang J., Chen Y., Wu Y, & Wang X. (2023). Regulation of Schwann cell proliferation and migration via miR-195-5p-induced Crebl2 downregulation upon peripheral nerve damage. Frontiers in Cellular Neuroscience, 17, 1173086.

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Neuromuscular Junction Morphology Analysis

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Following perfusion, the forelimb muscles (flexor digitorum profundus (FDP) and palmaris longus (PL)) were harvested, fixed in PFA, and cryoprotected in 30% sucrose for 24–48 h before embedding in Shandon M1 embedding matrix (Thermo Fisher Scientific; Waltham, MA). Transverse sections of 40 µm thickness were obtained and co-labeled with the NF200 antibody (1:200; Sigma Aldrich; St. Louis, MO) to visualize the heavy microfilament (200 kDa) in axons, and FITC-conjugated a-bungarotoxin (1:1000; Invitrogen; Carlsbad, CA) to visualize the nicotinic acetylcholine receptors (AChR). The images are presented with maximum intensity projection and the muscle fiber area was quantified by measuring the area of individual fibers using ImageJ.

Meyers E.C., Kasliwal N., Solorzano B.R., Lai E., Bendale G., Berry A., Ganzer P.D., Romero-Ortega M., Rennaker RL I.I., Kilgard M.P, & Hays S.A. (2019). Enhancing plasticity in central networks improves motor and sensory recovery after nerve damage. Nature Communications, 10, 5782.

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Quantifying Nerve Regeneration in Rats

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On day 28 after surgery, six rats from each group were randomly selected and sacrificed to harvest nerve grafts (n = 6). All nerves were fixed in 4% paraformaldehyde for 24 hours at 4°C, followed by dehydration in 20% sucrose solution for 24 hours and a second dehydration in 30% sucrose solution for an additional 24 hours. The middle portion of the nerve was cut into 10-μm-thick sections using a cryostat. These sections were then mounted on slides and subjected to immunohistochemistry with a rabbit anti-rat neurofilament 200 (NF-200) antibody (1:400; Sigma-Aldrich, Tokyo, Japan) and goat anti-rabbit secondary antibody conjugated to fluorescein isothiocyanate (1:200; TCS Biologicals, Botolph Clayton, UK). Antibodies were diluted in PBS solution containing 3% rat serum, 3% goat serum, and 0.02% sodium azide (Sigma-Aldrich) to reduce background staining.
Computer-assisted imaging analysis (Seescan Analytical Services, Cambridge, UK) was used to identify regenerating axons and blood vessels within grafts. The middle portions of grafts were examined. The integrated optical density of positive staining was assessed using Image-Pro Plus 6.0 (Media Cybernetics, Rockville, MD, USA). Three random sections were selected from each rat for analysis and the results were averaged.

Huang Y.Y., Xu X.L., Huang X.J., Liu J.H., Qi J., Zhu S., Zhu Z.W., He B., Zhu Q.T., Xu Y.B., Gu L.Q, & Liu X.L. (2018). Various changes in cryopreserved acellular nerve allografts at −80°C. Neural Regeneration Research, 13(9), 1643-1649.

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Neurite growth and cell death analysis

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Neuronal Neuro2a (N2a) and HeLa cells were cultures in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with glutamax, 1% penicillin-streptomycin, and 10% fetal calf serum (all Gibco). Cells were co-transfected with Fugene 6 (Promega) for 48 h as per the manufacturer’s protocol. shRNA constructs were from Sigma (#11857 for Gpi1 and as previously described for Oxr1 [14 (link)]). For neurite growth assay, medium was replaced with serum-free medium when transfecting and incubated for 48 h. For cell death assay, cells were treated with 250-μM arsenite (Sigma) for 4 h. Cells were fixed with 4% paraformaldehyde for 10 min at room temperature, washed with PBS (Sigma) twice, and blocked with blocking buffer (5% goat serum (VectorLab), 0.5% Triton X-100 (Sigma)) for 1 h at room temperature. Neurites and pyknotic nuclei were visualised with NF200 antibody (Sigma N4142) and DAPI staining (Vectorlabs), respectively.

Finelli M.J., Paramo T., Pires E., Ryan B.J., Wade-Martins R., Biggin P.C., McCullagh J, & Oliver P.L. (2018). Oxidation Resistance 1 Modulates Glycolytic Pathways in the Cerebellum via an Interaction with Glucose-6-Phosphate Isomerase. Molecular Neurobiology, 56(3), 1558-1577.

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Immunohistochemical Profiling of Neuronal Markers

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Immunohistochemistry was performed according to standard protocols, and the following primary antibodies were used: Isolectin B4 antibody (Biotinylated GRIFFONIA, 1:200, vector laboratories, B-1205), CGRP antibody (Goat, 1:300, Abcam, ab36001), NF200 antibody (mouse, 1:200, Sigma-Aldrich, N5389), CCR2 antibody (rabbit, 1:300, NOVOUS, NBP1-48,337), Alexa Fluor® 594 (donkey anti-rabbit IgG, 1:1000, Abcam, ab150132), Alexa Fluor® 488 (donkey anti-mouse IgG, 1:1000, Abcam, ab150105; donkey anti-goat IgG, 1:1000, Abcam, ab150129). Alexa Fluor 488-conjugated streptavidin to visualize the riboprobes (1:1000, Invitrogen, E13345).

Ma S.B., Xian H., Wu W.B., Ma S.Y., Liu Y.K., Liang Y.T., Guo H., Kang J.J., Liu Y.Y., Zhang H., Wu S.X., Luo C, & Xie R.G. (2020). CCL2 facilitates spinal synaptic transmission and pain via interaction with presynaptic CCR2 in spinal nociceptor terminals. Molecular Brain, 13, 161.

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Neurite Growth and Cell Death Assays

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