重组Anti-Ki67抗体[EPR3610] - BSA and Azide free (ab209897)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR3610] to Ki67 - BSA and Azide free
- Suitable for: Flow Cyt (Intra), IHC-P, WB, ICC/IF
- Knockout validated
- Reacts with: Human
Related conjugates and formulations
概述
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产品名称
Anti-Ki67抗体[EPR3610] - BSA and Azide free
参阅全部 Ki67 一抗 -
描述
兔单克隆抗体[EPR3610] to Ki67 - BSA and Azide free -
宿主
Rabbit -
经测试应用
适用于: Flow Cyt (Intra), IHC-P, WB, ICC/IFmore details -
种属反应性
与反应: Human
不与反应: Mouse, Rat -
免疫原
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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阳性对照
- WB: HeLa and ramos cell lysates. IHC-P: Human tonsil, colon, ovarian carcinoma, squamous cell carcinoma of cervix and colonic adenocarcinoma tissues. ICC/IF: HeLa, HT-29 cells, KI67-HAP1 cells (WT and KO). Flow Cyt (intra): Ramos cells, HAP1 cells.
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常规说明
ab209897 is the carrier-free version of ab92742.
This product is not suitable for xenograft experiments. For further information please contact our Customer Services team.
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.
This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.
Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.
This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
性能
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形式
Liquid -
存放说明
Shipped at 4°C. Store at +4°C. Do Not Freeze. -
存储溶液
pH: 7.20
Constituent: PBS -
无载体
是 -
Concentration information loading...
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纯度
Protein A purified -
克隆
单克隆 -
克隆编号
EPR3610 -
同种型
IgG -
研究领域
相关产品
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Alternative Versions
- Alexa Fluor® 647 Anti-Ki67 antibody [EPR3610] (ab196907)
- Alexa Fluor® 488 Anti-Ki67 antibody [EPR3610] (ab197234)
- Alexa Fluor® 568 Anti-Ki67 antibody [EPR3610] (ab211968)
- HRP Anti-Ki67 antibody [EPR3610] (ab212215)
- Alexa Fluor® 555 Anti-Ki67 antibody [EPR3610] (ab215226)
- Alexa Fluor® 594 Anti-Ki67 antibody [EPR3610] (ab216709)
- APC Anti-Ki67 antibody [EPR3610] (ab310886)
- PE Anti-Ki67 antibody [EPR3610] (ab310942)
- Anti-Ki67 antibody [EPR3610] (ab92742)
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Compatible Secondaries
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Conjugation kits
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Isotype control
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Positive Controls
应用
The Abpromise guarantee
Abpromise™承诺保证使用ab209897于以下的经测试应用
“应用说明”部分 下显示的仅为推荐的起始稀释度;实际最佳的稀释度/浓度应由使用者检定。
应用 | Ab评论 | 说明 |
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Flow Cyt (Intra) |
1/100 - 1/150.
ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
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IHC-P | (1) |
1/500 - 1/1000.
See IHC antigen retrieval protocols. For unpurified use at 1/500 - 1/1000. |
WB |
1/5000. Predicted molecular weight: 359 kDa.
For unpurified use at 1/500 - 1/1000. |
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ICC/IF |
Use a concentration of 1 µg/ml.
If fixing cells in 4% PFA, it is recommended to permeabilized cells with 0.1% Triton-X for 5 min. |
说明 |
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Flow Cyt (Intra)
1/100 - 1/150. ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
WB
1/5000. Predicted molecular weight: 359 kDa. For unpurified use at 1/500 - 1/1000. |
ICC/IF
Use a concentration of 1 µg/ml. If fixing cells in 4% PFA, it is recommended to permeabilized cells with 0.1% Triton-X for 5 min. |
靶标
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功能
Required to maintain individual mitotic chromosomes dispersed in the cytoplasm following nuclear envelope disassembly (PubMed:27362226). Associates with the surface of the mitotic chromosome, the perichromosomal layer, and covers a substantial fraction of the chromosome surface (PubMed:27362226). Prevents chromosomes from collapsing into a single chromatin mass by forming a steric and electrostatic charge barrier: the protein has a high net electrical charge and acts as a surfactant, dispersing chromosomes and enabling independent chromosome motility (PubMed:27362226). Binds DNA, with a preference for supercoiled DNA and AT-rich DNA (PubMed:10878551). Does not contribute to the internal structure of mitotic chromosomes (By similarity). May play a role in chromatin organization (PubMed:24867636). It is however unclear whether it plays a direct role in chromatin organization or whether it is an indirect consequence of its function in maintaining mitotic chromosomes dispersed. -
序列相似性
Contains 1 FHA domain.
Contains 16 K167R repeats.
Contains 1 PP1-binding domain. -
发展阶段
Expression occurs preferentially during late G1, S, G2 and M phases of the cell cycle, while in cells in G0 phase the antigen cannot be detected (at protein level) (PubMed:6206131). Present at highest level in G2 phase and during mitosis (at protein level). In interphase, forms fiber-like structures in fibrillarin-deficient regions surrounding nucleoli (PubMed:2674163, PubMed:8799815). -
翻译后修饰
Phosphorylated. Hyperphosphorylated in mitosis (PubMed:10502411, PubMed:10653604). Hyperphosphorylated form does not bind DNA. -
细胞定位
Chromosome. Nucleus. Nucleus, nucleolus. Associates with the surface of the mitotic chromosome, the perichromosomal layer, and covers a substantial fraction of the mitotic chromosome surface (PubMed:27362226). Associates with satellite DNA in G1 phase (PubMed:9510506). Binds tightly to chromatin in interphase, chromatin-binding decreases in mitosis when it associates with the surface of the condensed chromosomes (PubMed:15896774, PubMed:22002106). Predominantly localized in the G1 phase in the perinucleolar region, in the later phases it is also detected throughout the nuclear interior, being predominantly localized in the nuclear matrix (PubMed:22002106). - Information by UniProt
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数据库链接
- Entrez Gene: 4288 Human
- Omim: 176741 Human
- SwissProt: P46013 Human
- Unigene: 689823 Human
- Unigene: 80976 Human
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别名
- Antigen identified by monoclonal antibody Ki 67 antibody
- Antigen identified by monoclonal antibody Ki-67 antibody
- Antigen KI-67 antibody
see all
图片
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This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab92742).
Flow cytometry overlay histogram showing wild-type Hap1 (green line) and MKI67 knockout Hap1 stained with ab92742 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilised with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS containing 10% normal goat serum to block non-specific protein-protein interaction followed by the antibody (ab92742) (1x 106 in 100μl at 0.04 μg/ml (1/57000)) for 30min at 22°C.
The secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed was incubated at 1/4000 for 30min at 22°C
Isotype control antibody Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control was used at the same concentration and conditions as the primary antibody (wild-type Hap1 - black line, MKI67 knockout Hap1 - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity).
Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.
This antibody gave a positive signal in Hap1 Fixed with 4% formaldehyde (10 min) / permeabilised with 0.1% PBS-Triton X-100 for 15 min under the same conditions.
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Comparison between RNApII-S2P-/low cells and Ki-67- cells
a: Regulation of Ki-67 and RNApII-S2P during proliferation and quiescence in T98G glioblastoma cells. T98G cells were grown in culture medium containing 10% (v/v) fetal bovine serum (FBS), were induced to become quiescent by serum starvation in medium supplemented with 0.5% (v/v) FBS for 14 days, and then were re-stimulated by being split 1:5 into new medium containing 10% (v/v) FBS and cultured for 3 days. The cells were detached from dishes with trypsin-EDTA solution, fixed in 10% (v/v) neutral buffered formalin, and centrifuged. Paraffin sections of the pellet were cut, and expression of Ki-67 and RNApII-S2P was examined by single (brown; a1, a2, a4, a5, a7, a8) or double immunostaining (Ki-67, brown; RNApII-S2P, red; a3, a6, a9). Hematoxylin (blue) was used as a nuclear stain. Ki-67- RNApII-S2P-/low cells (blue cells in the double stained sections) emerged only in the quiescent condition (a6, arrows). Scale bar, 10 μm. b: Single-color immunostaining for Ki-67 (b1) and RNApII-S2P (b2) in serial sections of glioblastoma tissue. Ki-67- tumor cells were frequently found, whereas only a few RNApII-S2P-/low cells (arrows) were observed around necrotic area. N, necrotic area; V, blood vessels. Scale bars, 50 μm.
Ki67 detected using ab92742.
(From Figure S2 of Ishii et al)
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab92742).
Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
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Clone EPR3610 (ab209897) has been successfully conjugated by Abcam. This image was generated using Anti-Ki67 antibody [EPR3610] (Alexa Fluor® 488). Please refer to ab197234 for protocol details.
ab197234 staining Ki67 in wild-type HAP1 cells (top panel) and Ki67 knockout HAP1 cells (bottom panel). The cells were fixed with 100% methanol (5min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab197234 at 1/100 dilution (shown in green) and ab195889 at 1/250 dilution (shown in pseudo colour red) overnight at +4°C. Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
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Anti-Ki67 antibody [EPR3610] (ab92742) at 1/5000 dilution (purified) + Ramos (Human Burkitt's lymphoma cell line) cell lysate at 20 µg
Secondary
Peroxidase-conjugated goat anti-rabbit IgG, (H+L) at 1/1000 dilution
Predicted band size: 359 kDa
Observed band size: 395 kDa why is the actual band size different from the predicted?Blocking and dilution buffer: 5% NFDM/TBST.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab92742).
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Clone EPR3610 (ab209897) has been successfully conjugated by Abcam. This image was generated using Anti-Ki67 antibody [EPR3610] (Alexa Fluor® 647). Please refer to ab196907 for protocol details.
ab196907 staining Ki67 in wild-type HAP1 cells (top panel) and Ki67 knockout HAP1 cells (bottom panel). The cells were fixed with 100% methanol (5min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab196907 at 1/100 dilution (shown in red) and ab195887 at 1/250 dilution (shown in green) overnight at +4°C. Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
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Chromogenic triple immunostaining for estrogen receptor (ER), progesterone receptor (PgR), and Ki-67 in breast cancer tissue to verify the triple immunostaining detection method.
Panel e: ER+ PgR- Ki-67- cells were stained red (short arrow), ER- PgR+ Ki-67- cells were stained blue (black arrowhead), ER+ PgR+ Ki-67- cells were stained purple (long arrow), and Ki-67+ cells were stained brown (white arrowhead). These colors are easily distinguishable. Scale bars, 25 μm.
Deparaffinized sections were pretreated for antigen retrieval by boiling in antigen retrieval solution, pH 9. Sections were incubated with rabbit monoclonal antibody against Ki67 ab92742 at a 1/1000 dilution. After the reaction with (HRP)-conjugated secondary antibodies color was developed with (DAB) and sections were counterstained with hematoxylin.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab92742).
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ab92742 staining Ki67 in human adenocarcinoma cells by ICC/IF (Immunocytochemistry/immunofluorescence).
Cells were fixed with paraformaldehyde and permeabilized with 0.1% Triton X-100 in PBS and blocked with 5% serum for 1 hour at 21°C. Samples were incubated with primary antibody (1/1000) for 12 hours at 4°C. A Cy3® conjugated donkey anti-rabbit IgG polyclonal was used as the secondary antibody at a dilution of 1/200.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab92742).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human squamous cell carcinoma of cervix tissue labeling Ki67 with purified ab92742 at 1/250. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Counterstained with hematoxylin.
Negative control using PBS instead of primary antibody (inset).
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab92742).
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Anti-Ki67 antibody [EPR3610] (ab92742) at 1/500 dilution (unpurified) + HeLa (Human epithelial cell line from cervix adenocarcinoma) cell lysate at 10 µg
Secondary
HRP-conjugated goat anti-rabbit IgG at 1/2000 dilution
Predicted band size: 359 kDa
Observed band size: 395 kDa why is the actual band size different from the predicted?This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab92742).
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Immunocytochemistry/Immunofluorescence analysis of HT-29 (Human colorectal adenocarcinoma cell line) cells labeling Ki67 with purified ab92742 at 1/250. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain. ab7291, a mouse anti-tubulin (1/1000) and ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/500) were also used.
Control 1: primary antibody (1/250) and secondary antibody, ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/500).
Control 2: ab7291 (1/1000) and secondary antibody, ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/500).
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab92742).
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Intracellular Flow Cytometry analysis of Ramos (Human Burkitt's lymphoma cell line) cells lablling Ki67 with purified ab92742 at 1/150 (red). Cells were fixed with 2% paraformaldehyde. An FITC-conjugated goat anti-rabbit IgG (1/150) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabeled control, cells without incubation with primary and secondary antibodies.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab92742).
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ab92742 staining Ki67 in wild-type HAP1 cells (top panel) and Ki67 knockout HAP1 cells (bottom panel). The cells were fixed with 100% methanol (5min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab92742 at 1µg/ml and ab195889 at 1/250 dilution (shown in pseudo colour red) overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labeled in blue with DAPI.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab92742).
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Overlay histogram showing Ramos (Human Burkitt's lymphoma cell line) cells stained with unpurified ab92742 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (unpurified ab92742, 1/100 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1µg/1x106 cells) used under the same conditions.
Acquisition of >5,000 events was performed.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab92742).
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Immunocytochemistry/Immunofluorescence analysis of HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling Ki67 with unpurified ab92742 at a dilution of 1/250.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab92742).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human colonic adenocarcinoma tissue labeling Ki67 with unpurified ab92742.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab92742).
Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human normal colon tissue labeling Ki67 with unpurified ab92742.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab92742).
Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human ovarian carcinoma tissue labeling Ki67 with unpurified ab92742.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab92742).
Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human cervical carcinoma tissue labeling Ki67 with unpurified ab92742.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab92742).
Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
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ab209897 staining Ki67 in wild-type HAP1 cells (top panel) and Ki67 knockout HAP1 cells (bottom panel). The cells were fixed with 4% formaldehyde (10min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab209897 at 1μg/ml concentration and ab195889 at 1/250 dilution (shown in pseudo colour red) overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.
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Immunohistochemical analysis of paraffin-embedded Human tonsil tissue labeling Ki67 with ab209897 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.
Nucleus staining on human tonsil.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.
Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
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Immunohistochemical analysis of paraffin-embedded Human bladder cancer labeling Ki67 with ab209897 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.
Nucleus staining on human bladderr cancer.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.
Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
实验方案
数据表及文件
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Datasheet download
Certificate of Compliance
文献 (10)
ab209897 被引用在 10 文献中.
- Zheng Y et al. PD-L1+CD8+ T cells enrichment in lung cancer exerted regulatory function and tumor-promoting tolerance. iScience 25:103785 (2022). Mass Cytometry, glucose transporter glut1 . PubMed: 35146396
- Cho WC et al. Exosomal miR-193a and let-7g accelerate cancer progression on primary colorectal cancer and paired peritoneal metastatic cancer. Transl Oncol 14:101000 (2021). PubMed: 33352502
- Yan Y et al. PRMT5 regulates colorectal cancer cell growth and EMT via EGFR/Akt/GSK3ß signaling cascades. Aging (Albany NY) 13:4468-4481 (2021). PubMed: 33495409
- Ehteda A et al. Dual targeting of the epigenome via FACT complex and histone deacetylase is a potent treatment strategy for DIPG. Cell Rep 35:108994 (2021). PubMed: 33852836
- Li S et al. MicroRNA-4500 Inhibits Migration, Invasion, and Angiogenesis of Breast Cancer Cells via RRM2-Dependent MAPK Signaling Pathway. Mol Ther Nucleic Acids 21:278-289 (2020). PubMed: 32615527
- Wang Y et al. Upregulated Circular RNA circ-UBE2D2 Predicts Poor Prognosis and Promotes Breast Cancer Progression by Sponging miR-1236 and miR-1287. Transl Oncol 12:1305-1313 (2019). PubMed: 31336316
- He Q et al. Methionine Sulfoxide Reductase B1 Regulates Hepatocellular Carcinoma Cell Proliferation and Invasion via the Mitogen-Activated Protein Kinase Pathway and Epithelial-Mesenchymal Transition. Oxid Med Cell Longev 2018:5287971 (2018). Human . PubMed: 29861830
- Zhang J et al. SOX4 inhibits GBM cell growth and induces G0/G1 cell cycle arrest through Akt-p53 axis. BMC Neurol 14:207 (2014). WB ; Human . PubMed: 25366337
- Brassart-Pasco S et al. Tetrastatin, the NC1 domain of the a4(IV) collagen chain: a novel potent anti-tumor matrikine. PLoS One 7:e29587 (2012). IHC-Fr . PubMed: 22539938
- Chen Z et al. Clinicopathological significance of non-small cell lung cancer with high prevalence of Oct-4 tumor cells. J Exp Clin Cancer Res 31:10 (2012). IHC-P ; Human . PubMed: 22300949