DYKDDDDK Tag Antikörper

Produktnummer ABIN559688

Produktdetails anti-DYKDDDDK Tag Antikörper

Target: DYKDDDDK Tag

Reaktivität: Bitte anfragen

Wirt: Maus

Klonalität: Monoklonal

Konjugat: Dieser DYKDDDDK Tag Antikörper ist unkonjugiert

Applikation: Western Blotting (WB), ELISA, Immunoprecipitation (IP), Dot Blot (DB), Immunostaining (ISt)

Sequenz: DYKDDDDK

Spezifität: Recognizes the N-terminal, C-terminal or internal DYKDDDDK-tagged fusion proteins

Sensitivität: WB (with ECL): 1-5ng of DYKDDDDK-tagged fusion proteins

Aufreinigung: Protein A affinity chromatography from mouse ascites fluid

Immunogen: DYKDDDDK (DYKDDDDK) synthetic peptide conjugated to KLH

Klon: FG4R

Isotyp: IgG1

Anwendungsinformationen

Applikationshinweise: WB (with ECL): 1:1000-3000 dilution (incubate for one hour at room temperature) / IS: 1:500-2000 dilution / For best results with other assays (e.g.: Dot, ELISA, IP, etc), please determine optimal working dilution by titration test

Beschränkungen: Nur für Forschungszwecke einsetzbar

Independent Validation (IF)

Validierung #102796 (Immunofluorescence)

by: Martinelli lab, Neuroscience Department, UConn Health

No.: #102796

Datum: 13.02.2018

Antigen: DYKDDDDK-tag

Chargennummer: 02017

Validierte Anwendung: Immunofluorescence

Positivkontrolle: African green monkey kidney fibroblast-like COS7 cells transfected with BAI3, a transmembrane protein with a surface presented DYKDDDDK-tag

Negativkontrolle: untransfected COS7 cells

Bewertung:

Passed. Signal was observed as expected. There was never any signal on untransfected cells, demonstrating specificity of the antibody.

Validierungsbilder

COS7 cells were transfected to express a DYKDDDDK-tagged and surface presented BAI3 protein. Indirect immunoflouresence using ABIN1715174 reveals specific signal for the DYKDDDDK-tag (middle) on the cell surface. No colocalization of the ABIN1715174 and nuclear stain (DAPI, left) was observed (merge, right). No signal was ever observed in any untransfected cell.

Protokoll

Primärantikörper: ABIN1715174

Sekundärantikörper: goat anti-mouse AF488 conjugated antibody (Life Technologies, A11029, lot 1829920)

Full Protocol:

• Grow COS7 cells in DMEM (Biological Industries, 01-052-1A, lot 1728619) supplemented with FBS (Hyclone, SH3039602) without antibiotics, at 37°C and 5% CO₂ in 1ml on glass coverslips on 24-well plate.
• Transfect cells with calcium phosphate method with a eukaryotic expression construct to express a DYKDDDDK-tagged BAI3, which will be surface presented.
• Grow cells for 24h.
• Fix cells on coverslips in 4% PFA for 15min at 4°C.
• Wash cells 3x for 5min with PBS.
• Block non-specific binding with 5% goat serum in PBS for 1h at RT.
• Incubate cells with primary mouse anti-DYKDDDDK-tag antibody (antibodies-online, ABIN1715174, lot 02017) diluted 1:2000 in blocking solution ON at 4°C.
• Wash cells 3x for 5min with PBS.
• Incubate cells with secondary goat anti-mouse AF488 conjugated antibody (Life Technologies, A11029, lot 1829920) diluted 1:400 in blocking buffer for 1h at RT protected from light.
• Wash cells 3x for 5min with PBS.
• Mount coverslips on glass slides in Fluoromount-G with DAPI mounting medium (ThermoFisher Scientific, 00-4959-52, lot B2215-N915).
• Image acquisition on a Zeiss Axiovert epifluorescence with with a 40x objective, using a Zeiss ApoTome for optical sectioning.

Handhabung

Konzentration: 1mg/mL

Buffer: 10mM PBS, pH 7.2, 0.05% NaN3

Konservierungsmittel: Sodium azide

Vorsichtsmaßnahmen: This product contains sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.

Lagerung: -20 °C

Details zu DYKDDDDK Tag

Target: DYKDDDDK Tag

Andere Bezeichnung: FLAG Tag (DYKDDDDK Tag Produkte)

Substanzklasse: Tag