Afadin, also MLLT-4 (Myeloid/Lymphoid or mixed-lineage Leukemia Translocated to Ch 4) and AF6, is a 200-210 kDa nectin and F-actin-binding intracellular protein. It is ubiquitously expressed, and serves as an intermediary that links nectin with the actin cytoskeleton in tight and adherens junctions (AJs). Afadin is involved in the nectin-mediated recruitment of catenins to zonule adherens complexes, and in the recruitment of ZO-1 to the apical side of AJs for the purpose of forming tight junctions. Human Afadin is 1824 amino acids (aa) in length. The protein contains two Ras-association (RA) domains (aa 20-348), a phosphoprotein FHA domain (aa 426‑492), a Vav2 Ras-activating "dilute" domain (aa 668-908), one PDZ domain that binds protein C-termini (aa 1007-1093), and three proline-rich regions (aa 1346‑1708). There are at least fourteen utilized Ser/Thr phosphorylation sites and two Tyr phosphorylation sites. There are at least seven alternative splice variants are reported for Afadin. There is a 190 kDa isoform that shows a 12 aa insert after Arg1605 coupled to a four aa substitution for aa 1650-1824. Other isoforms contain scattered deletions of one to three aa coupled to major aa changes. Five isoforms share a deletion of aa 393-407, with four of these containing a 24 aa substitution for aa 1605-1824, a 73 aa substitution for aa 1747-1824, an 11 aa substitution for aa 680-1824, and a deletion of aa 1683-1746, respectively. A final isoform possesses a four aa substitution for aa 1650-1824. Over aa 217-392, human Afadin shares 94% aa sequence identity with mouse Afadin.
Human Afadin/AF-6 Antibody
R&D Systems | Catalog # AF7829
Key Product Details
Species Reactivity
Validated:
Human
Cited:
Mouse
Applications
Validated:
Immunohistochemistry, Western Blot, Immunocytochemistry
Cited:
Immunohistochemistry
Label
Unconjugated
Antibody Source
Polyclonal Sheep IgG
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Product Specifications
Immunogen
E. coli-derived recombinant human Afadin/AF-6
Asn217-Pro392
Accession # P55196
Asn217-Pro392
Accession # P55196
Specificity
Detects human Afadin/AF-6 in direct ELISAs and Western blots. In direct ELISAs, less than 1% cross-reactivity with recombinant human METTL-11A is observed.
Clonality
Polyclonal
Host
Sheep
Isotype
IgG
Scientific Data Images for Human Afadin/AF-6 Antibody
Detection of Human Afadin/AF-6 by Western Blot.
Western blot shows lysates of HEK293T human embryonic kidney cell line and SK-OV-3 human ovarian adenocarcinoma cell line. PVDF membrane was probed with 1 µg/mL of Sheep Anti-Human Afadin/AF-6 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7829) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). A specific band was detected for Afadin/AF-6 at approximately 220 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.Afadin/AF-6 in BEAS2B Human Cell Line.
Afadin/AF-6 was detected in immersion fixed BEAS2B human lung epithelial cell line using Sheep Anti-Human Afadin/AF-6 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7829) at 1.7 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Sheep IgG Secondary Antibody (red; Catalog # NL010) and counterstained with DAPI (blue). Specific staining was localized to plasma membrane. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.Afadin/AF-6 in Human Brain.
Afadin/AF-6 was detected in immersion fixed paraffin-embedded sections of human brain (hippocampus) using Sheep Anti-Human Afadin/AF-6 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7829) at 3 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Sheep HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS019) and counterstained with hematoxylin (blue). Specific staining was localized to neuronal cell bodies and processes. View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.Applications for Human Afadin/AF-6 Antibody
Application
Recommended Usage
Immunocytochemistry
1-15 µg/mL
Sample: Immersion fixed BEAS2B human lung epithelial cell line
Sample: Immersion fixed BEAS2B human lung epithelial cell line
Immunohistochemistry
3-15 µg/mL
Sample: Immersion fixed paraffin-embedded sections of human brain (hippocampus)
Sample: Immersion fixed paraffin-embedded sections of human brain (hippocampus)
Western Blot
1 µg/mL
Sample: HEK293T human embryonic kidney cell line and SK‑OV‑3 human ovarian adenocarcinoma cell line
Sample: HEK293T human embryonic kidney cell line and SK‑OV‑3 human ovarian adenocarcinoma cell line
Formulation, Preparation, and Storage
Purification
Antigen Affinity-purified
Reconstitution
Sterile PBS to a final concentration of 0.2 mg/mL. For liquid material, refer to CoA for concentration.
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Formulation
Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.
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.
Calculators
Background: Afadin/AF-6
Alternate Names
AF-6, AF6, AFAD, MLLT4
Gene Symbol
AFDN
UniProt
Additional Afadin/AF-6 Products
Product Documents for Human Afadin/AF-6 Antibody
Certificate of Analysis
To download a Certificate of Analysis, please enter a lot or batch number in the search box below.
Note: Certificate of Analysis not available for kit components.
Product Specific Notices for Human Afadin/AF-6 Antibody
For research use only
Related Research Areas
Citations for Human Afadin/AF-6 Antibody
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Protocols
Find general support by application which include: protocols, troubleshooting, illustrated assays, videos and webinars.
- Antigen Retrieval Protocol (PIER)
- Antigen Retrieval for Frozen Sections Protocol
- Appropriate Fixation of IHC/ICC Samples
- Cellular Response to Hypoxia Protocols
- Chromogenic IHC Staining of Formalin-Fixed Paraffin-Embedded (FFPE) Tissue Protocol
- Chromogenic Immunohistochemistry Staining of Frozen Tissue
- ClariTSA™ Fluorophore Kits
- Detection & Visualization of Antibody Binding
- Fluorescent IHC Staining of Frozen Tissue Protocol
- Graphic Protocol for Heat-induced Epitope Retrieval
- Graphic Protocol for the Preparation and Fluorescent IHC Staining of Frozen Tissue Sections
- Graphic Protocol for the Preparation and Fluorescent IHC Staining of Paraffin-embedded Tissue Sections
- Graphic Protocol for the Preparation of Gelatin-coated Slides for Histological Tissue Sections
- ICC Cell Smear Protocol for Suspension Cells
- ICC Immunocytochemistry Protocol Videos
- ICC for Adherent Cells
- IHC Sample Preparation (Frozen sections vs Paraffin)
- Immunocytochemistry (ICC) Protocol
- Immunocytochemistry Troubleshooting
- Immunofluorescence of Organoids Embedded in Cultrex Basement Membrane Extract
- Immunofluorescent IHC Staining of Formalin-Fixed Paraffin-Embedded (FFPE) Tissue Protocol
- Immunohistochemistry (IHC) and Immunocytochemistry (ICC) Protocols
- Immunohistochemistry Frozen Troubleshooting
- Immunohistochemistry Paraffin Troubleshooting
- Preparing Samples for IHC/ICC Experiments
- Preventing Non-Specific Staining (Non-Specific Binding)
- Primary Antibody Selection & Optimization
- Protocol for Heat-Induced Epitope Retrieval (HIER)
- Protocol for Making a 4% Formaldehyde Solution in PBS
- Protocol for VisUCyte™ HRP Polymer Detection Reagent
- Protocol for the Fluorescent ICC Staining of Cell Smears - Graphic
- Protocol for the Fluorescent ICC Staining of Cultured Cells on Coverslips - Graphic
- Protocol for the Preparation & Fixation of Cells on Coverslips
- Protocol for the Preparation and Chromogenic IHC Staining of Frozen Tissue Sections
- Protocol for the Preparation and Chromogenic IHC Staining of Frozen Tissue Sections - Graphic
- Protocol for the Preparation and Chromogenic IHC Staining of Paraffin-embedded Tissue Sections
- Protocol for the Preparation and Chromogenic IHC Staining of Paraffin-embedded Tissue Sections - Graphic
- Protocol for the Preparation and Fluorescent ICC Staining of Cells on Coverslips
- Protocol for the Preparation and Fluorescent ICC Staining of Non-adherent Cells
- Protocol for the Preparation and Fluorescent ICC Staining of Stem Cells on Coverslips
- Protocol for the Preparation and Fluorescent IHC Staining of Frozen Tissue Sections
- Protocol for the Preparation and Fluorescent IHC Staining of Paraffin-embedded Tissue Sections
- Protocol for the Preparation of Gelatin-coated Slides for Histological Tissue Sections
- Protocol for the Preparation of a Cell Smear for Non-adherent Cell ICC - Graphic
- R&D Systems Quality Control Western Blot Protocol
- TUNEL and Active Caspase-3 Detection by IHC/ICC Protocol
- The Importance of IHC/ICC Controls
- Troubleshooting Guide: Immunohistochemistry
- Troubleshooting Guide: Western Blot Figures
- Western Blot Conditions
- Western Blot Protocol
- Western Blot Protocol for Cell Lysates
- Western Blot Troubleshooting
- Western Blot Troubleshooting Guide
- View all Protocols, Troubleshooting, Illustrated assays and Webinars
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