重组Anti-Aurora B抗体[EP1009Y]

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Aurora B antibody [EP1009Y] (AB45145)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human endometrium carcinoma tissue sections labeling Aurora B with purified ab45145 at 1/200 dilution (1.35 µg/mL). Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0). The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Negative control : PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-Aurora B antibody [EP1009Y] (AB45145)

Intracellular Flow Cytometry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling Aurora B with Purified ab45145 at 1/300 dilution (0.1 μg/ml) (Red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor^® 488 ,ab150077) secondary antibody was used at 1/2000. Isotype control - Rabbit monoclonal IgG (Left). Unlabeled control - /.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Aurora B antibody [EP1009Y] (AB45145)

Immunocytochemistry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling Aurora B with Purified ab45145 at 1 : 50 dilution (5.4 µg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) 1 : 200 (2.5 µg/ml). Goat anti rabbit IgG (Alexa Fluor® 488,ab150077) was used as the secondary antibody at 1 : 1000 (2 µg/ml) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

• IP

Lab

Immunoprecipitation - Anti-Aurora B antibody [EP1009Y] (AB45145)

Purified ab45145 at 1/20 dilution (1μg) immunoprecipitating Aurora B in HeLa whole cell lysate.
Lane 1 (input) : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate 10μg
Lane 2 (+) : ab45145 + HeLa whole cell lysate.
Lane 3 (-) : Rabbit monoclonal IgG (ab172730) instead of ab45145 in HeLa whole cell lysate.
VeriBlot for IP Detection Reagent (HRP) (ab131366) (1/1000 dilution) was used for Western blotting.
Blocking Buffer and concentration : 5% NFDM/TBST.
Diluting buffer and concentration : 5% NFDM/TBST.
Observed band size : 39 kDa

All lanes:

Immunoprecipitation - Anti-Aurora B antibody [EP1009Y] (ab45145)

Predicted band size: 39 kDa

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• WB

Lab

Western blot - Anti-Aurora B antibody [EP1009Y] (AB45145)

This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 2% Bovine Serum Albumin before being incubated with ab45145 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution ab133406

Lanes 1 - 2:

Western blot - Anti-Aurora B antibody [EP1009Y] (ab45145) at 1/1000 dilution

Lanes 3 - 4:

Western blot - Anti-Aurora B antibody [EP1009Y] (ab45145) at 1/5000 dilution

Lanes 5 - 6:

Western blot - Anti-Aurora B antibody [EP1009Y] (ab45145) at 1/10000 dilution

Lanes 7 - 8:

Western blot - Anti-Aurora B antibody [EP1009Y] (ab45145) at 1/50000 dilution

Lanes 9 - 10:

Western blot - Anti-Aurora B antibody [EP1009Y] (ab45145) at 1/75000 dilution

Lanes 1, 3, 5, 7 and 9:

HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate at 10 µg

Lanes 2, 4, 6, 8 and 10:

HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate Nocodozole Stimulated at 10 µg

Secondary

All lanes:

Western blot - VHH Single Domain Anti-Rabbit IgG Fc (HRP) (<a href='/products/secondary-antibodies/vhh-single-domain-rabbit-igg-fc-hrp-ab191866'>ab191866</a>) at 1/10000 dilution

Predicted band size: 39 kDa

Observed band size: 39 kDa

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Exposure time: 8min

• OI-RD Scanning

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OI-RD Scanning - Anti-Aurora B antibody [EP1009Y] (AB45145)

We have systematically measured KD (the equilibrium dissociation constant between the antibody and its antigen), of more than 840 recombinant antibodies to assess not only their individual KD values but also to see the average affinity of antibody. Based on the comparison with published literature values for mouse monoclonal antibodies, Recombinant antibodies appear to be on average 1-2 order of magnitude higher affinity.

• WB

CiteAb

Western blot - Anti-Aurora B antibody [EP1009Y] (AB45145)

Aurora B western blot using anti-Aurora B antibody [EP1009Y] ab45145. Publication image and figure legend from Chuang, M. J., Wu, S. T., et al., 2013, PLoS One, PubMed 24023871.

ab45145 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab45145 please see the product overview.

HDACI induced G2/M phase cell cycle and growth inhibition in prostate cancer cells via distinct mechanisms.(A) LBH589 had a dose-dependent effect on growth inhibition in prostate cancer cell lines. The cell lines were treated with 50, 75, 100, and 150 nM of LBH589. Cell survival was measured using MTT assay as described in the Materials and Methods section. Graphs compared the percentage of growth inhibition to vehicle control at 24 (upper) and 48 (lower) hours. (B) LBH589 induced G2/M cell cycle arrest. All cell lines were treated with LBH589 or vehicle control for 24 h. The cell cycles were analyzed by PI-staining and flow cytometry according to DNA content. The increased percentages of G2/M cells were compared to that of vehicle control. (C) LBH589 induced apoptosis in PC-3 and LNCaP cells. Both cell lines were treated with 75nM LBH589 or vehicle control for 24 (upper) and 48 (lower) h. The percentages of apoptotic cells were analyzed by counting the population of DNA fragmentation by flow cytometry. (D–E) HDACIs induced Aurora kinase degradation found in PC-3 but not in LNCaP. PC-3 and LNCaP cells were treated with different HDACIs at indicated concentrations of LBH589, SAHA and SBHA for 24 h and cell lysates were analyzed by western blot using the indicated antibodies. (F) Profile of proteins regarding progression of G2/M cell cycle. The cells were treated with different dosages of LBH589 for 24 h. The lysate was analyzed by western blot.

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