Isolated neurons were washed with PBS, permeabilized with 4% formaldehyde and 0.5% Triton X-100 for 30 min in 4°C. Cells were washed in PBS twice, blocked in 5% bovine serum albumin (Merck Millipore, Darmstadt, Germany) for 1 h in 4°C, and incubated with neurofilament antibody (1:500, cat. no. ab8135, Abcam, Cambridge, UK) at 4°C overnight. Following a wash with PBS, the cells were stained with a FITC-conjugated secondary antibody (1:2,000, cat. no. ab6717, Abcam) for 1 h at room temperature. Then, cells were captured using intravital upright microscope (ECLIPSEFN1; Nikon Corporation, Tokyo, Japan). Results indicated that the purity of neuron preparations was >90%, as required for onward experiments.
Neurofilament antibody
Manufactured by Merck Group
Sourced in United States
The Neurofilament antibody is a laboratory reagent used to detect and quantify neurofilament proteins, which are structural components of neurons. It can be used in various analytical techniques, such as Western blotting and immunohistochemistry, to study the expression and distribution of neurofilaments in biological samples.
Automatically generated - may contain errors
Lab products found in correlation
Bovine serum albumin (bsa), by Merck Group (1 mentions)
Eclipse fn1, by Nikon (1 mentions)
Dmem f12, by Thermo Fisher Scientific (1 mentions)
Map2 antibody, by Merck Group (1 mentions)
Microplate reader, by Thermo Fisher Scientific (1 mentions)
Nestin antibody, by Abcam (1 mentions)
Epidermal growth factor (egf), by Thermo Fisher Scientific (1 mentions)
Basic fibroblast growth factor (bfgf), by Thermo Fisher Scientific (1 mentions)
Trypsin, by Merck Group (1 mentions)
Cold centrifuge, by Merck Group (1 mentions)
β catenin antibody, by Santa Cruz Biotechnology (1 mentions)
Gsk3β antibody, by Santa Cruz Biotechnology (1 mentions)
Alexa fluor 594, by Thermo Fisher Scientific (1 mentions)
Prolong gold antifade reagent with dapi, by Thermo Fisher Scientific (1 mentions)
Fv1000 confocal laser scanning microscope, by Olympus (1 mentions)
Anti iba1 antibody, by Fujifilm (1 mentions)
Darpp 32 antibody, by Abcam (1 mentions)
Smi 99p, by Fortrea (1 mentions)
4 protocols using neurofilament antibody
1
Immunostaining of Isolated Neurons
2
Stem Cell Differentiation Assay
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The following drugs and reagents were used: lithium chloride (Alfa Aesar B21573, Haverhill, MA, USA), β-catenin antibody (Santa Cruz C2206, Dallas, TX, USA), GSK-3β antibody (Santa Cruz AB15328), Nestin antibody (Abcam ab6142, Cambridgeshire, UK), MAP2 antibody (Merck Millipore AB5622, Burlington, MA, USA), GFAP antibody (Merck Millipore MAB3402C3), neurofilament antibody (Merck Millipore AB15328, MA), DMEM/F12 (Thermo Fisher, Waltham, MA, USA), basic fibroblast growth factor (bFGF, PeproTech, Rocky Hill, NJ, USA), epidermal growth factor (EGF, PeproTech), 0.25% trypsin (Sigma Aldrich, St. Louis, MO, USA), 0.01 mol/L RNase (Sigma Aldrich), 0.5 mg/L propidium iodide staining solution (Boster Biology, Wuhan, China), 5% Chloral hydrate (China), and 4% paraformaldehyde (Boster Biology). The equipment included a cold centrifuge (Sigma Aldrich, USA), an inverted light microscope (Olympus IX70, Tokyo, Japan), a microplate reader (Thermo Fisher), an incubator that was set at a temperature of 37°C and regulated with 5% CO2 (Thermo Fisher), and flow cytometer (FACSCalibur 2, Becton Dickinson, Franklin Lakes, NJ, USA).
3
Neurofilament Immunohistochemistry of Optic Nerve
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Optic nerves were isolated, processed, and embedded in paraffin (n = 2). 5 μm longitudinal paraffin sections of the optic nerve were deparaffinized and rehydrated, followed by incubation with blocking buffer (1% BSA, 0.1% Triton X-100, and 1% normal donkey serum in 1 X PBS) in a humidified chamber for 1 h at RT. The slides were then incubated overnight at 4°C with a Neurofilament antibody (Sigma-Aldrich, catalog #: MAB5266, St. Louis, MO, USA). The next day, samples were washed with 1X PBS three times for 5 min each, followed by incubation with anti-mouse Alexa Fluor 594 (ThermoFisher Scientific, catalog #: A11020, Waltham, MA, USA), staining was performed for 1 h at RT. The coverslips were then mounted on slides using proLong Gold antifade reagent with DAPI (4′,6-diamidino-2-phenylindole) (Invitrogen, catalog #: P36935, Waltham, MA, USA). Slides were observed under an Olympus FV1000 Confocal laser scanning microscope, and images were captured using 60X objective with the use of appropriate filters and lasers.
4
Quantitative Analysis of Brain Proteins
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