SUN5 Antikörper
Kurzübersicht für SUN5 Antikörper (ABIN1876373)
Target
Alle SUN5 Antikörper anzeigenReaktivität
Wirt
Klonalität
Konjugat
Applikation
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Produktmerkmale
- Expected molecular weight of target: 379 kDa
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Aufreinigung
- Affinity purification
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Isotyp
- IgG
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Applikationshinweise
- WB 1:500-1:2000
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Beschränkungen
- Nur für Forschungszwecke einsetzbar
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- by
- Johann-Friedrich-Blumenbach-Institute for Zoology and Anthropology, Department of Developmental Biology, Georg-August-University Göttingen
- No.
- #102344
- Datum
- 19.12.2017
- Antigen
- SUN5
- Chargennummer
- Validierte Anwendung
- Immunocytochemistry
- Positivkontrolle
- NIH/3T3 mouse embryonic fibroblast cells overexpressing HA- and myc-tagged SPAG4L-2/SUN5
- Negativkontrolle
- NIH/3T3 mouse embryonic fibroblast cells overexpressing HA- and myc-tagged SPAG4
- Bewertung
Passed. ABIN1876373 is specific for SPAG4L-2/SUN5 and did not detect SPAG4 when ectopically expressed. Unspecific binding to other cellular proteins is nearly absent.
- Primärantikörper
- ABIN1876373
- Sekundärantikörper
- goat anti-rabbit IgG DyLight488 (Thermo Scientific, 35553, lot KA130088)
- Full Protocol
- NIH/3T3 cells (ATCC, CRL-1658) are grown in DMEM+GlutaMAX (Gibco, 31966-021, Lot 1852045) supplemented with fetal bovine serum (Gibco 270-106) and Pen/Strep (Gibco 15140), at 37°C and 5% CO2 to 70% confluency.
- Transfect cells with HA/myc-tagged SPAG4 expression plasmid or HA/myc-tagged SPAG4L-2/SUN5 (Frohnert et al., 2011) expression plasmid using EndofectinMax (GeneCopoeia) following the manufacturer's instructions.
- Grow cells for 24h.
- Fix cells in 3.7% paraformaldehyde (in PBS) for 15min at 4°C followed by incubation in 0.3% Triton X-100 for 10min at 4°C.
- Block cells in PBS containing 1% bovine serum albumin and 0.5% Tween-20 (PBT) for 1h at room temperature.
- Incubate cells with primary
- rabbit anti-SUN5 antibody (antibodies-online, ABIN1876373) diluted 1:100 in PBS ON at 4° and
- mouse anti-myc tag antibody clone (9E10) diluted 1:100 in PBS ON at 4°C.
- Wash cells with TBST (50mM Tris-HCl, pH 7.4, 150mM NaCl, 0.1% Tween 20) for 15min at RT.
- Incubate cells with secondary
- goat anti-rabbit IgG DyLight488 (Thermo Scientific, 35553, lot KA130088) diluted 1:1000 in PBS for 1h at 37°C and
- goat anti-mouse IgG (H+L) Alexa Fluor 555 (Invitrogen, A21422, lot 948498) diluted 1:1000 in PBS for 1h at 37°C.
- Counterstain DNA with DAPI (4,6-Diamidino-2-phenylindole; Sigma D-9542).
- Image acquisition on Zeiss LSM 510 confocal microscope and processing using Adobe Photoshop 5.0.
- Anmerkungen
Validierung #102344 (Immunocytochemistry)ValidierungsbilderSpecificity of ABIN1876373. Cells were transfected either with SPAG4 (upper row) or SPAG4L-2/SUN5 (lower row), both tagged with HA and myc. Anti-myc antibody (B, F; red) detected both proteins, but ABIN1876373 (A, E; green) exclusively detected SPAG4L-2/SUN5 without any cross-reaction to SPAG4. The antibody detected SPAG4L-2/SUN5 at the nuclear membrane, as expected. The pictures on the right (D, H) show the red and green channels merged with DAPI counterstain (blue).
Protokoll -
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Format
- Liquid
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Konzentration
- 1 mg/mL
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Buffer
- PBS with 0.02 % sodium azide, 50 % glycerol, pH 7.4
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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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Handhabung
- Avoid repeated freeze/thaw cycles.
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Lagerung
- -20 °C
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Informationen zur Lagerung
- Store at -20°C.
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- SUN5 (Sad1 and UNC84 Domain Containing 5 (SUN5))
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Andere Bezeichnung
- SPAG4L
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Gen-ID
- 140732
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UniProt
- Q8TC36
Target
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