BrdU Antikörper

Produktnummer ABIN2477713

Produktdetails anti-BrdU Antikörper

Target: BrdU (Bromodeoxyuridine (BrdU))

Reaktivität: Bitte anfragen

Wirt: Maus

Klonalität: Monoklonal

Konjugat: Dieser BrdU Antikörper ist unkonjugiert

Applikation: Flow Cytometry (FACS), Immunohistochemistry (Paraffin-embedded Sections) (IHC (p)), Immunocytochemistry (ICC), Immunohistochemistry (Frozen Sections) (IHC (fro))

Produktmerkmale: Mouse anti BrdU antibody, clone Bu20a, recognizes BrdU and other thymidine analogs; 5′-chloro-2′-deoxyuridine (CldU), 5′-iodo-2′-deoxyuridine (IdU) and 2′-deoxy-5-ethynyluridine (EdU), but only shows minimal reactivity with thymidine itself

Aufreinigung: Purified IgG prepared by affinity chromatography on Protein G from tissue culture supernatant

Immunogen: Bromodeoxyuridine conjugated to BSA

Klon: Bu20a

Isotyp: IgG1

Anwendungsinformationen

Applikationshinweise: Flow: 1/25-1/100, ICC 1/25-1/100
For Flow Cytometry use 10 μl of the suggested working dilution to label 1x106 cells in 100 μl

Beschränkungen: Nur für Forschungszwecke einsetzbar

Independent Validation (IHC)

Validierung #103189 (Immunohistochemistry)

by: Prof. Merighi, Laboratory of Neurobiology, Department of Veterinary Sciences, University of Turin

No.: #103189

Datum: 05.10.2018

Antigen: BrdU

Chargennummer: 1710

Validierte Anwendung: Immunohistochemistry

Positivkontrolle:

Adult (24 months) mouse brain and intestine (duodenum)

Negativkontrolle:

We incubated slices overnight with the blocking solution only and then processed them with the secondary antibody.

Bewertung:

Passed. ABIN2477713 works well in IHC-P, but with higher concentrations than the ones suggested by the manufacturer.

Validierungsbilder

Staining of mouse brain and intestine sections using ABIN2477713. A-C: A bright staining in the cell nuclei of individual cells scattered across the brain is present, especially in the hippocampus, ventricular wall (A) and sub-cortical areas at the border between the gray and white matters (B-C). Some positive cells are clearly M-phase cells (C). Abbreviations: GM = gray matter; LV = lateral ventricle; WM = white matter. D-E: positive control staining in the intestine (duodenum). BrdU+ proliferating cells are clearly detected in the germinal layer of the intestinal epithelium, at the basis of the duodenal villi.

Protokoll

Primärantikörper: ABIN2477713

Sekundärantikörper: goat anti-mouse AF488 conjugated antibody (Thermo Fisher Scientific, A11029, lot 1942237)

Full Protocol:

• Inject mouse intraperitoneally at the dose of 0.1 mg/g body weight with BrdU (Sigma, B5002) dissolved in sterile distilled water at final concentration of 100mg/ml.
• Perfuse mouse was perfused with paraformaldehyde 4% in 0.1M phosphate buffer (PB) pH 7.4 and post-fix samples in the same fixative for an additional 2h at RT.
• Wash, dehydrate, and embed samples in paraffin wax.
• Following several washes in PBS, cut intestines and brain with a microtome (6µm-thick sections) and mount sections on glass slides.
• After paraffin removal, process sections by microwave antigen retrieval for 10min (95-100°C) in 10mM sodium citrate buffer (pH6.0).
• Let sections cool for 20min.
• Wash sections 2x 5min with distilled water.
• Wash sections 5min with PBS.
• Incubate sections in PBS containing 1% albumin from chicken egg white (Sigma, A5378) and 0.3% Triton-X-100 (BioRad, 161-0407, lot 00583) for 1h at RT to block non-specific binding sites.
• Incubate sections with primary antibody mouse anti-BrdU (antibodies-online, ABIN2477713, lot 1710) diluted 1:10 in PBS-BSA-PLL ON at RT.
• Wash sections 3x 5min with 0.01M PBS.
• Incubate sections with the secondary goat anti-mouse AF488 conjugated antibody (Thermo Fisher Scientific, A11029, lot 1942237) diluted 1:200 in 0.1M PBS for 1h at RT.
• Wash sections 3x 5min with 0.01M PBS.
• Mount sections in Fluoroshield (Sigma, F6182, lot MKCB0153V).
• Acquire images with Leica DM 6000B fluorescence microscope equipped with the manufacturer’s FITC filter set and digital camera at magnifications of 20x and 40x. Parameters for image acquisition (Exposure 2.2s, Gain 2.3x, Saturation 1.5, Gamma 1.10) were maintained unchanged for all images.

Anmerkungen:

For the incubation with the primary antibody ABIN2477713 1:25, 1:20, 1:15 dilutions were also tested with also 0.01M PBS - 5% Normal Goat Serum (NGS Sigma, G9023, lot SLBV1396) and 0.1% Triton-X -100 as the blocking solution, but 1:10 and PBS containing 1% albumin from chicken egg white and 0.3% Triton-X gave the best results.

Handhabung

Format: Liquid

Konzentration: 1.0 mg/mL

Buffer: Phosphate buffered saline, 0.09%Sodium Azide (NaN3)

Konservierungsmittel: Sodium azide

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

Handhabung: This product should be stored undiluted. Avoid repeated freezing and thawing as this may denature the antibody. Should this product contain a precipitate we recommend microcentrifugation before use.

Lagerung: 4 °C/-20 °C

Informationen zur Lagerung: Store at +4oC or at -20oC if preferred. Storage in frost-free freezers is not recommended.

Haltbarkeit: 18 months

Referenzen für anti-BrdU Antikörper (ABIN2477713)

Innis, Dai, Wu, Buchan, Jacobson: "Perinatal lipid nutrition alters early intestinal development and programs the response to experimental colitis in young adult rats." in: American journal of physiology. Gastrointestinal and liver physiology, Vol. 299, Issue 6, pp. G1376-85, (2010) (PubMed).

Details zu BrdU

Target: BrdU (Bromodeoxyuridine (BrdU))

Andere Bezeichnung: BrdU (BrdU Produkte)

Substanzklasse: Chemical

Hintergrund: Synonyms: 5-BROMODEOXYURIDINE