DYKDDDDK Tag Antikörper
Kurzübersicht für DYKDDDDK Tag Antikörper (ABIN559688)
Target
Reaktivität
Wirt
Klonalität
Konjugat
Applikation
Klon
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Sequenz
- DYKDDDDK
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Spezifität
- Recognizes the N-terminal, C-terminal or internal DYKDDDDK-tagged fusion proteins
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Sensitivität
- WB (with ECL): 1-5ng of DYKDDDDK-tagged fusion proteins
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Aufreinigung
- Protein A affinity chromatography from mouse ascites fluid
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Immunogen
- DYKDDDDK (DYKDDDDK) synthetic peptide conjugated to KLH
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Isotyp
- IgG1
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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
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Beschränkungen
- Nur für Forschungszwecke einsetzbar
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- 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.
- 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.
- Anmerkungen
Validierung #102796 (Immunofluorescence)ValidierungsbilderCOS7 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 -
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Konzentration
- 1mg/mL
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Buffer
- 10mM PBS, pH 7.2, 0.05% NaN3
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Konservierungsmittel
- Sodium azide
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Vorsichtsmaßnahmen
- This product contains sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.
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Lagerung
- -20 °C
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- DYKDDDDK Tag
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Andere Bezeichnung
- FLAG Tag
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Substanzklasse
- Tag
Target
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