Biolegend, 608451, Purified anti-RBFOX3 (NeuN) Antibody, 25μg

RBFox3, commonly referred to as NeuN, is one of a family of 3 mammalian Fox homologues. Fox was discovered in C. elegans as a gene involved in sex determination, and the name RBFox is an acronym of "Feminizing locus on X". The Fox protein and its 3 mammalian homologues are all about 46 kD proteins, each of which includes a central highly conserved RRM type RNA recognition motif, which corresponds to a small ~70 amino acid structure consisting of 4 beta strands and two alpha-helices. RBFox3 is a protein which has a function in RNA splicing and is expressed heavily and specifically in neuronal nuclei. The antibody reveals strong nuclear and distal cytoplasmic staining for RBFox3/NeuN and the complete absence of staining of astrocytes and other kinds of non-neuronal cells. This RBFox3/NeuN antibody is therefore an excellent marker of neuronal cells.
25μg
Verified Reactivity: Human, Mouse
Antibody Type: Monoclonal
Host Species: Rat
Immunogen: Full length recombinant human RBFOX3, isoform 5
Formulation: Phosphate-buffered solution, pH 7.2, containing 0.09% sodium azide
Preparation: The antibody was purified by affinity chromatography.
Concentration: 0.5 mg/mL
Storage & Handling: The antibody solution should be stored undiluted between 2°C and 8°C.
Application: WB - Quality tested IHC-P, IHC-F - Verified SB - Community verified
Recommended Usage: Each lot of this antibody is quality control tested by western blotting. For western blotting, the suggested use of this reagent is 0.125 - 1.0 µg/mL. For immunohistochemistry on formalin-fixed paraffin-embedded tissue sections, a concentration range of 1.0 - 10.0 µg/mL is suggested. For immunohistochemistry on frozen tissue sections, a concentration range of 1.0 - 10.0 µg/mL is suggested. It is recommended that the reagent be titrated for optimal performance for each application.
Additional Product Notes: This product has been verified for IHC-F (Immunohistochemistry - frozen tissue sections) on the NanoString GeoMx® Digital Spatial Profiler. The GeoMx® enables researchers to perform spatial analysis of protein and RNA targets in FFPE and fresh frozen human and mouse samples. For more information about our spatial biology products and the GeoMx® platform, please visit our spatial biology page.
RRID: AB_2927957 (BioLegend Cat. No. 608451) AB_2927957 (BioLegend Cat. No. 608452)
Structure: RBFox3 is a 312 amino acid protein with a predicted molecular weight of 34 kD
Distribution: Mature Neurons, Neural Stem Cells/ Cytoplasm and Nucleus
Function: Cell Biology, Neuroscience, Neuroscience Cell Markers, Stem Cells
Cell Type: Mature Neurons, Neural Stem Cells
Biology Area: Cell Biology, Neuroscience, Neuroscience Cell Markers, Stem Cells
Molecular Family: Nuclear Markers
Antigen References: Dhekne HS, et al. 2018. Elife. 7:e40202. Sun Y, et al. 2020. J Neuroinflammation. 17:72. Schwabenland M, et al. 2021. Immunity. 54:1594-1610.
Gene ID: 146713
UniProt: View information about RBFOX3 on UniProt.org
Clone: W18230A
Regulatory Status: RUO
Other Names: NeuN, RNA binding protein fox-1 homolog 3 (RBFox3), hexaribonucleotide binding protein 3, HRNBP3
Isotype: Rat IgG2a, κ
Q: If an antibody clone has been previously successfully used in IBEX in one fluorescent format, will other antibody formats work as well?
A: It’s likely that other fluorophore conjugates to the same antibody clone will also be compatible with IBEX using the same sample fixation procedure. Ultimately a directly conjugated antibody’s utility in fluorescent imaging and IBEX may be specific to the sample and microscope being used in the experiment. Some antibody clone conjugates may perform better than others due to performance differences in non-specific binding, fluorophore brightness, and other biochemical properties unique to that conjugate.
Q: Will antibodies my lab is already using for fluorescent or chromogenic IHC work in IBEX?
A: Fundamentally, IBEX as a technique that works much in the same way as single antibody panels or single marker IF/IHC. If you’re already successfully using an antibody clone on a sample of interest, it is likely that clone will have utility in IBEX. It is expected some optimization and testing of different antibody fluorophore conjugates will be required to find a suitable format; however, legacy microscopy techniques like chromogenic IHC on fixed or frozen tissue is an excellent place to start looking for useful antibodies.
Q: Are other fluorophores compatible with IBEX?
A: Over 18 fluorescent formats have been screened for use in IBEX, however, it is likely that other fluorophores are able to be rapidly bleached in IBEX. If a fluorophore format is already suitable for your imaging platform it can be tested for compatibility in IBEX.
Q: The same antibody works in one tissue type but not another. What is happening?
A: Differences in tissue properties may impact both the ability of an antibody to bind its target specifically and impact the ability of a specific fluorophore conjugate to overcome the background fluorescent signal in a given tissue. Secondary stains, as well as testing multiple fluorescent conjugates of the same clone, may help to troubleshoot challenging targets or tissues. Using a reference control tissue may also give confidence in the specificity of your staining.
Q: How can I be sure the staining I’m seeing in my tissue is real?
A: In general, best practices for validating an antibody in traditional chromogenic or fluorescent IHC are applicable to IBEX. Please reference the Nature Methods review on antibody based multiplexed imaging for resources on validating antibodies for IBEX.