Anti-BrdU 抗体 [BU1/75 (ICR1)] - Proliferation Marker (ab6326)
Key features and details
- Rat monoclonal [BU1/75 (ICR1)] to BrdU - Proliferation Marker
- Suitable for: ICC/IF, IHC-P, Flow Cyt
- Isotype: IgG2a
製品の概要
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製品名
Anti-BrdU antibody [BU1/75 (ICR1)] - Proliferation Marker
BrdU 一次抗体 製品一覧 -
製品の詳細
Rat monoclonal [BU1/75 (ICR1)] to BrdU - Proliferation Marker -
由来種
Rat -
アプリケーション
適用あり: ICC/IF, IHC-P, Flow Cytmore details -
種交差性
Not applicable. -
免疫原
The details of the immunogen for this antibody are not available.
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ポジティブ・コントロール
- ICC/IF: HeLa cells; Flow Cyt: HeLa cells; IHC-P: Rat small intestine tissue.
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特記事項
Protocol advice:
This antibody recognizes single stranded DNA so the DNA needs to be unraveled first. This can be done with DNAse, although it doesn't give the best results. Depending on the assay, acid denaturation with 2M HCL or heat denaturation are the most successful. Please note this step is critical in any assay with this antibody and is the area that should be modified to optimize results. A detailed BrdU staining protocol is available in the Protocols tab or by clicking on this link.
Unstained positive control slides from mice treated with BrdU (formalin-fixed, paraffin-embedded intestine sections) are available as BrdU control slides ab129956.
This monoclonal antibody is manufactured exclusively by Abcam.
AF488 conjugate available as ab220074
AF647 conjugate available as ab220075
Please see the Associated Products tab for other available conjugates.If you require this antibody in a particular buffer formulation or a particular conjugate for your experiments, please contact orders@abcam.com or you can find further information here.
Reproducibility is key to advancing scientific discovery and accelerating scientists’ next breakthrough.
Abcam is leading the way with our range of recombinant antibodies, knockout-validated antibodies and knockout cell lines, all of which support improved reproducibility.
We are also planning to innovate the way in which we present recommended applications and species on our product datasheets, so that only applications & species that have been tested in our own labs, our suppliers or by selected trusted collaborators are covered by our Abpromise™ guarantee.
In preparation for this, we have started to update the applications & species that this product is Abpromise guaranteed for.
We are also updating the applications & species that this product has been “predicted to work with,” however this information is not covered by our Abpromise guarantee.
Applications & species from publications and Abreviews that have not been tested in our own labs or in those of our suppliers are not covered by the Abpromise guarantee.
Please check that this product meets your needs before purchasing. If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, as well as customer reviews and Q&As.
製品の特性
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製品の状態
Liquid -
保存方法
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle. -
バッファー
pH: 7.40
Preservative: 0.02% Sodium azide
Constituent: PBS
Some batches contain 6.97% L-Arginine as a stabilizing agent. For lot-specific buffer information, please contact our Scientific Support team. -
Concentration information loading...
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精製度
IgG fraction -
ポリ/モノ
モノクローナル -
クローン名
BU1/75 (ICR1) -
アイソタイプ
IgG2a -
軽鎖の種類
kappa -
研究分野
関連製品
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Alternative Versions
- Alexa Fluor® 488 Anti-BrdU antibody [BU1/75 (ICR1)] (ab220074)
- Alexa Fluor® 647 Anti-BrdU antibody [BU1/75 (ICR1)] (ab220075)
- Alexa Fluor® 594 Anti-BrdU antibody [BU1/75 (ICR1)] (ab220076)
- HRP Anti-BrdU antibody [BU1/75 (ICR1)] (ab220507)
- Alexa Fluor® 555 Anti-BrdU antibody [BU1/75 (ICR1)] (ab221240)
- Anti-BrdU antibody [BU1/75 (ICR1)] - BSA and Azide free (ab264079)
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Compatible Secondaries
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Isotype control
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Related Products
アプリケーション
Our Abpromise guarantee covers the use of ab6326 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
アプリケーション | Abreviews | 特記事項 |
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ICC/IF | 1/250. | |
IHC-P | Use a concentration of 1 - 3 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol. | |
Flow Cyt | 1/25 - 1/200. Rat IgG2a, kappa monoclonal [RTK2758] (Low endotoxin, Azide Free) (ab18450), is suitable for use as an isotype control with this antibody. |
ターゲット情報
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関連性
The immunocytochemical detection of bromodeoxyuridine (BrdU) incorporated into DNA is a powerful tool to study the cytokinetics of normal and neoplastic cells. In vitro or in vivo labeling of tumor cells with the thymidine analogue BrdU and the subsequent detection of incorporated BrdU with specific anti-BrdU monoclonal antibodies is an accurate and comprehensive method to quantitate the degree of DNA-synthesis. BrdU is incorporated into the newly synthezised DNA of S-phase cells may provide an estimate for the fraction of cells in S-phase. Also dynamic proliferative information such as the S-phase transit rate and the potential doubling time can be obtained, by means of bivariate BrdU/DNA flow cytometric analysis. -
細胞内局在
Nuclear -
別名
- Bromodeoxyuridine antibody
- BUdr antibody
画像
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Immunocytochemistry/ Immunofluorescence - Anti-BrdU antibody [BU1/75 (ICR1)] - Proliferation Marker (ab6326)
ab6326 stained in Hela cells. Untreated and BrdU treated (10uM for 24 hours) cells were fixed with 100% methanol (5 min) and then subjected to acid hydrolysis using 2M HCL in 0.1% PBS-Tween for 30 minutes at room temperature to denature the DNA. They were then incubated with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% triton for 1h at room temperature to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab6326 at 5µg/ml and ab7291 (Mouse monoclonal to alpha tubulin) at 1ug/ml overnight at +4°C. The secondary antibodies were Goat Anti-Rat IgG H&L (Alexa Fluor® 488) preadsorbed (ab150165) (colored green) used at 2 ug/ml and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (ab150120) (pseudo-colored red) used at 1/1000 dilution for 1hour at room temperature. DAPI was used to stain the cell nuclei (colored blue) at a concentration of 1.43µM for 1hour at room temperature.
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Dot plot showing BrdU-treated HeLa cells stained with ab6326. Cells were incubated with 10 µM BrdU for 30 minutes prior to being harvested, washed twice in 1x PBS and fixed in 70% ethanol (4°C, added drop-wise) for at least 30 minutes on ice. Once fixed, pellets were acid denatured with 2M HCl for 30 minutes at 22ºC and then neutralised with borate buffer (0.1M, pH8.5).
Samples were washed and incubated in 1x PBS / 10% normal goat serum to block non-specific protein-protein interactions followed by the antibody (ab6326, 1µg/1x106 cells) for 30 min at 22°C. The secondary antibody used was Goat Anti-Rat IgG H&L (Alexa Fluor® 488) preadsorbed (ab150165) at 1/2000 dilution for 30 min at 22ºC.
7-AAD was added to cells 20 min prior to data acquisition.
Acquisition of >5,000 events were collected using a 50 mW Blue laser (488nm) with 530/30 and 685/35 bandpass filters.
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Immunocytochemistry/ Immunofluorescence - Anti-BrdU antibody [BU1/75 (ICR1)] - Proliferation Marker (ab6326)Image from Condezo GN et al., PLoS Pathog. 2017;13(4):e1006320. Fig 1.; doi: 10.1371/journal.ppat.1006320. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/
Confocal immunofluorescence sections (~0.3 μm thick) showing the localization of the different factors in HEK 293 cells infected with Ad5 wt. Double labeling for BrdU (ab6326, magenta) and L1 52/55 kDa (green). Dotted rectangles indicate the areas shown at higher magnification in the bottom row. Bars: 3 μm.
Monolayer HEK293 cells grown in cover glasses were infected with Ad5 wt. The infection was synchronized by incubating the cells for 30 min at 4°C and then 30 min at 37°C. Then, the inoculums were removed and medium was added. For BrdU labeling, after 18 h at 37°C the medium was changed by medium containing 25 μg/ml BrdU (5-Bromo-2’-deoxyuridine), followed by another change at 25 hpi. Incubation with BrdU proceeded at 37°C. After 36 hpi, the medium was removed and 4% paraformaldehyde in PBS was added to the cells during 10 min. After 3 rinses with PBS, cover glasses were incubated with a mixture of 0.5% saponin and 10% FBS in PBS for 10 min. Samples were incubated with the primary antibody in 0.5% saponin and 2% FBS in PBS during 45 min. After three more rinses, incubation with secondary antibodies diluted in 0.5% saponin and 2% FBS in PBS was carried out in darkness. After a final rinse with PBS, cover glasses were mounted on glass slides. Antifade reagent was allowed to dry overnight before sample observation. All incubations were carried out at room temperature. Images were taken using a confocal multispectral Leica TCS SP5 system.
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Immunocytochemistry/ Immunofluorescence - Anti-BrdU antibody [BU1/75 (ICR1)] - Proliferation Marker (ab6326)
ICC/IF image of ab6326 stained HeLa cells, both BrdU treated (left image) and normal cells (right image). The cells were 100% methanol fixed (5 min) and then subjected to acid hydrolysis using 2M HCL in 0.1% PBS-Tween for 30 minutes at room temperature to denature the DNA. They were then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab6326, 10µg/ml) overnight at +4°C. The secondary antibody (green) was Goat Anti-Rat IgG H&L (DyLight® 488) preadsorbed (ab98420) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM. Positive staining can be seen in the BrdU treated cells, but not in the normal cells, demonstrating specificity for BrdU.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BrdU antibody [BU1/75 (ICR1)] - Proliferation Marker (ab6326)
IHC image of ab6326 staining in a formalin-fixed, paraffin-embedded rat small intestine BrdU tissue section. The section was pre-treated using pressure cooker heat mediated antigen retrieval with sodium citrate buffer (pH6) for 30mins, and incubated overnight at +4°C with ab6326 at 3 ug/ml. A goat anti-rat biotinylated secondary antibody was used to detect the primary, and visualized using an HRP conjugated ABC system. The section was counterstained with haematoxylin and mounted with DPX.
For other IHC staining systems (automated and non-automated) users should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BrdU antibody [BU1/75 (ICR1)] - Proliferation Marker (ab6326)This image is courtesy of an anonymous Abreview
ab6326 staining cultured cells of rat brain tissue by ICC. The sample was PFA fixed and permeabilized in 1M HCl prior to blocking with 5% serum for 1 hour at 25°C. The primary antibody was diluted 1/500 and incubated with the sample for 16 hours at 25°C. A biotinylated rabbit anti-rat IgG antibody, diluted 1/200, was used as the secondary.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BrdU antibody [BU1/75 (ICR1)] - Proliferation Marker (ab6326)Image from Vandenplas S et al., PLoS One. 2016;11(4):e0152870. Fig 2.; doi: 10.1371/journal.pone.0152870. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/
Immunohistological staining of frontal (A-C) and sagittal (D) sections through a tooth family on the lower jaw of S. salar with BrdU labelled cells in green and DAPI counterstain in blue. The tooth families presented are in a similar state of development: a young functional tooth and its successor in morphogenesis stage. (A) Pulsed specimen, T0; (B) Chase time T1, one week after BrdU administration; (C) Chase time T2, two weeks after BrdU administration and (D) Chase time T3, four weeks after BrdU administration. The white arrow indicates single cells within the middle dental epithelium (MDE) enclosed by the IDE, cervical loop and ODE of the replacement tooth. Abbreviations: Ab: aboral; Ant: anterior; CV: cervical loop; DP: dental papilla; FT: functional tooth; IDE: inner dental epithelium; Lab: labial; Lin: lingual; MDE: middle dental epithelium; ME: mesenchyme; OE: oral epithelium; ODE: outer dental epithelium; Or: oral; PC: pulp cavity; Pos: posterior; yellow asterisk: replacement tooth; scale bars: 100 μm.
Immunohistological staining for BrdU on the paraffin sections: Rehydration through a decreasing ethanol series, chromatin precipitation in hydrochloric acid, block in 3% BSA/ 1% milk powder, exposure to primary antibody (ab6326) and secondary antibody (polyclonal anti-rat Alexa Fluor® 488), DAPI counterstaining (1μl/ml). Immunofluorescence was visualized on a NIKON eclips TE2000-S confocal laser-scanning microscope. Adobe Illustrator CS5, Adobe Photoshop CS5 and Fiji were used to process bright field and fluorescent images of the immunostained sections.
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Flow Cytometry - Anti-BrdU antibody [BU1/75 (ICR1)] - Proliferation Marker (ab6326)This image is courtesy of an anonymous Abreviewab6326 staining BrdU in HeLa cells by Flow Cytometry. Cells were incubated with 10 µM BrdU for 30 minutes prior to being harvested with 1X trypsin-EDTA, washed twice in PBS containing 1% BSA, and fixed in 70% ethanol (added drop-wise) for at least 30 minutes on ice. Once fixed, pellets were acid denatured with HCl/Triton X-100 for 30 minutes at room temperature and then neutralised with sodium tetraborate.
Pelleted cells were re-suspended in Tween/BSA/PBS to which primary antibody was then added (0.1 µg in 0.5% Tween 20 (v/v) plus 1% BSA in PBSA) and incubated for 30 minutes at room temperature. Secondary Alexa Fluor®488-conjugated Goat anti-Rat IgG (H+L) was used at 1/500 and incubated for 30 minutes at room temperature in the dark. Cells were pelleted once more and resuspended in PBS containing 5 µg/mL propidium iodide.
Gating Strategy: Based on forward and side scatter, cells were gated into the region used for analysis. This was done by applying a large circle to a -
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BrdU antibody [BU1/75 (ICR1)] - Proliferation Marker (ab6326)This image is courtesy of an anonymous Abreview
ab6326 staining BrdU in mouse lung tissue sections by Immunohistochemistry (Formalin/PFA-fixed parafin-embedded sections). Tissue samples were heat mediated with Citrate buffer for 40min. The sections were blocked with 10% donkey serum for 1 hour at 22°C. Samples were incubated with the primary antibody (1/200 in 10% Donkey serum-PBS 0.2% Triton) at 4°C for 16 hours. Cy™3 AffiniPure Donkey Anti-Rat IgG (H+L) (1/400) was used as the secondary antibody.
Representative confocal images of terminal bronchioles in mouse lungs taken at day 5 following naphthalene injection.
Colors label CC10 (red), FoxJ1 (green), BrdU (yellow), and nuclear DNA (blue).
参考文献 (1165)
ab6326 は 1165 報の論文で使用されています。
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