Human RBFOX3/NeuN MAb (Clone 1094645)

Catalog #: MAB11665 Datasheet / COA / SDS

Catalog #	Availability	Size / Price	Qty
MAB11665-100
MAB11665-SP

Detection of RBFOX3 in Human Cerebellum via seqIF™ staining on COMET™

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Human RBFOX3/NeuN MAb (Clone 1094645) Summary

Species Reactivity

Human

Specificity

Detects a synthetic peptide specific for human RBFOX3 around amino acid 5 in Direct ELISA.

Source

Monoclonal Mouse IgG₁ Clone # 1094645

Purification

Protein A or G purified from hybridoma culture supernatant

Immunogen

Synthetic Peptide
Accession # A6NFN3

Formulation

Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose.

Label

Unconjugated

Applications

Recommended Concentration

Sample

Western Blot

2 µg/mL

Human hippocampus tissue

Immunohistochemistry

3-25 µg/mL

Immersion fixed paraffin-embedded sections of human cerebellum

Multiplex Immunofluorescence

25 µg/mL

Immersion fixed paraffin-embedded sections of human cerebellum

Please Note: Optimal dilutions should be determined by each laboratory for each application. General Protocols are available in the Technical Information section on our website.

Scientific Data

Multiplex Immunofluorescence

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Detection of RBFOX3 in Human Cerebellum via seqIF™ staining on COMET™ RBFOX3 was detected in immersion fixed paraffin-embedded sections of human Cerebellum using Mouse Anti-Human RBFOX3, Monoclonal Antibody (Catalog #MAB11665) at 25ug/mL at 37 ° Celsius for 8 minutes. Before incubation with the primary antibody, tissue underwent an all-in-one dewaxing and antigen retrieval preprocessing using PreTreatment Module (PT Module) and Dewax and HIER Buffer H (pH 9; Epredia Catalog # TA-999-DHBH). Tissue was stained using the Alexa Fluor™ 647 Goat anti-Mouse IgG Secondary Antibody at 1:200 at 37 ° Celsius for 8 minutes. (Yellow; Lunaphore Catalog # DR647MS) and counterstained with DAPI (blue; Lunaphore Catalog # DR100). Specific staining was localized to the neuronal nuclei. Protocol available in COMET™ Panel Builder.

Immunohistochemistry

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Detection of RBFOX3/NeuN in Human Cerebellum. RBFOX3/NeuN was detected in immersion fixed paraffin-embedded sections of human cerebellum using Mouse Anti-Human RBFOX3/NeuN Monoclonal Antibody (Catalog # MAB11665) at 5 µg/ml for 1 hour at room temperature followed by incubation with the Anti-Mouse IgG VisUCyte™ HRP Polymer Antibody (Catalog # VC001) or the HRP-conjugated Anti-Mouse IgG Secondary Antibody (Catalog # HAF007). Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using VisUCyte Antigen Retrieval Reagent-Basic (Catalog # VCTS021). Tissue was stained using DAB (brown) and counterstained with hematoxylin (blue). Specific staining was localized to the nucleus. View our protocol for IHC Staining with VisUCyte HRP Polymer Detection Reagents.

Western Blot

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Detection of Human RBFOX3/NeuN by Western Blot. Western Blot shows lysates of human hippocampus tissue. PVDF membrane was probed with 2 µg/ml of Mouse Anti-Human RBFOX3/NeuN Monoclonal Antibody (Catalog # MAB11665) followed by HRP-conjugated Anti-Mouse IgG Secondary Antibody (Catalog # HAF018). Specific bands were detected for RBFOX3/NeuN at approximately 40 and 50 kDa (as indicated). This experiment was conducted under reducing conditions and using Western Blot Buffer Group 1.

Reconstitution Calculator

Preparation and Storage

Reconstitution

Reconstitute lyophilized material at 0.2mg/ml in sterile PBS. For liquid material, refer to CoA for concentration.

Shipping

Lyophilized product is shipped at ambient temperature. Liquid small pack size (-SP) is shipped with polar packs. Upon receipt, store immediately at the temperature recommended below.

Stability & Storage

Use a manual defrost freezer and avoid repeated freeze-thaw cycles.

12 months from date of receipt, -20 to -70 °C as supplied.
1 month, 2 to 8 °C under sterile conditions after reconstitution.
6 months, -20 to -70 °C under sterile conditions after reconstitution.

Background: RBFOX3/NeuN

RBFOX3, also known as Neuronal nuclei (NeuN), is a 34kDa marker of post-mitotic neurons that is highly conserved among different species. The RBFOX family regulates alternative slicing and is encoded by three genes: RBFOX1, RBFOX2 and RBFOX3. RBFOX3 mediates hippocampal circuitry, neurogenesis, and synaptogenesis. RBFOX3 plays a crucial role in normal synaptic function and is implicated in human neurological functions and mutations have been linked to neurodevelopmental delay, cognitive impairment, autistic features, and epilepsy.

References

Wang HY, Hsieh PF, Huang DF, Chin PS, Chou CH, Tung CC, Chen SY, Lee LJ, Gau SS, Huang HS. RBFOX3/NeuN is Required for Hippocampal Circuit Balance and Function. Sci Rep. 2015 Dec 1;5:17383. doi: 10.1038/srep17383. PMID: 26619789; PMCID: PMC4664964.
Huang DF, Lee CY, Chou MY, Yang TY, Cao X, Hsiao YH, Wu RN, Lien CC, Huang YS, Huang HP, Gau SS, Huang HS. Neuronal splicing regulator RBFOX3 mediates seizures via regulating Vamp1 expression preferentially in NPY-expressing GABAergic neurons. Proc Natl Acad Sci U S A. 2022 Aug 16;119(33):e2203632119. doi: 10.1073/pnas.2203632119. Epub 2022 Aug 11. PMID: 35951651; PMCID: PMC9388145.

Long Name

RNA binding fox-1 homolog 3

Entrez Gene IDs

146713 (Human); 52897 (Mouse); 287847 (Rat)

Alternate Names

FLJ56884; FLJ58356; Fox-1 homolog C; FOX3; FOX-3; hexaribonucleotide binding protein 3; HRNBP3; neuN antigen; NEUN; neuronal nuclei antigen; neuronal nuclei; RBFOX3; RNA binding protein fox-1 homolog 3; RNA binding protein, fox-1 homolog (C. elegans) 3; RNA binding protein, fox-1 homolog 3