Anti-Vimentin antibody [V9] - Cytoskeleton Marker

• 产品名称

  Anti-Vimentin抗体[V9] - Cytoskeleton Marker
  参阅全部 Vimentin 一抗

• 描述

  小鼠单克隆抗体[V9] to Vimentin - Cytoskeleton Marker

• 宿主

  Mouse

• 经测试应用

  适用于: ICC/IF, IHC-P, WB, Flow Cyt (Intra)more details

• 种属反应性

  与反应: Rat, Human
  预测可用于: Horse, Chicken, Cow, Cat, Dog, Pig

• 免疫原

  Full length native protein (purified). This information is proprietary to Abcam and/or its suppliers.

• 阳性对照

• WB: A549, Hap1, HeLa, U-2 OS, human tonsil and MOLT4 whole cell lysates. ICC/IF: HeLa, Hap1, SKOV-3 and rat glial cells. IHC-P: Human kidney and oral cavity tissue sections. Flow Cyt (Intra): HAP1, MDA-MB-231 and SV40LT-SMC cells.

• 常规说明

  This monoclonal antibody to vimentin has been knockout validated in ICC/IF. The expected staining was observed in wild type cells and no staining was seen in vimentin knockout cells. 

  
  

  This antibody clone is manufactured by Abcam. If you require a custom buffer formulation or conjugation for your experiments, please contact orders@abcam.com.

  
  

  The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing this with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation. 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, along with publications, customer reviews and Q&As

• 形式

  Liquid

• 存放说明

  Shipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid freeze / thaw cycles.

• 存储溶液

  pH: 7.40
  Preservative: 0.02% Sodium azide
  Constituents: PBS, 6.97% L-Arginine

• 浓度

• 10 µg 浓度为 1 mg/ml

• 100 µg 浓度为 1 mg/ml

• 纯度

  Affinity purified

• 克隆

  单克隆

• 克隆编号

  V9

• 同种型

  IgG1

The Abpromise guarantee

Abpromise™承诺保证使用ab8069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。

数据库链接

• Entrez Gene: 420519 Chicken

• Entrez Gene: 280955 Cow

• Entrez Gene: 477991 Dog

• Entrez Gene: 7431 Human

• Entrez Gene: 100522394 Pig

• Entrez Gene: 81818 Rat

• Omim: 193060 Human

• SwissProt: P09654 Chicken

• SwissProt: P48616 Cow

• SwissProt: P08670 Human

• SwissProt: P02543 Pig

• SwissProt: P31000 Rat

• Unigene: 455493 Human

• Unigene: 691131 Human

• Unigene: 2710 Rat

• 形式

  Vimentin is found in connective tissue and in the cytoskeleton.

• 别名

• HEL113 antibody

• VIM antibody

• VIME_HUMAN antibody

• Vimentin antibody

• CTRCT30 antibody

• Epididymis luminal protein 113 antibody

• FLJ36605 antibody

• 

  Western blot - Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069)

  All lanes : Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069) at 1 µg/ml
  
  Lane 1 : U-2 OS (Human bone osteosarcoma epithelial cell line) cell lysate
  Lane 2 : Human tonsil cell lysate
  Lane 3 : Wild-type HeLa cell lysate
  Lane 4 : VIM knockout HeLa cell lysate
  
  Lysates/proteins at 20 µg per lane.
  
  Performed under reducing conditions.
  
  Predicted band size: 54 kDa
  
  
  
  

  Lanes 1 - 4: Merged signal (red and green). Green - ab8069 observed at 53 kDa. Red - loading control, ab181602 observed at 37 kDa.

   ab8069 was shown to react with Vimentin in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab255446 (knockout cell lysate ab263775) was used. Wild-type and Vimentin knockout samples were subjected to SDS-PAGE. ab8069 and Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) were incubated overnight at 4°C at 1 µg/ml and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) and Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

   

   

• 

  Immunocytochemistry/ Immunofluorescence - Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069)

  ab8069 staining Vimentin (colored green) in wild-type HAP1 cells (top panel) and VIM 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 ab8069 at 0.5μg/ml and ab202272 (Rabbit monoclonal [EP1332Y] to alpha Tubulin (Alexa Fluor® 594)) at 1/250 dilution overnight at +4°C, followed by a further incubation at room temperature for 1h with ab150117 (Goat secondary antibody to Mouse IgG (Alexa Fluor® 488)) at 2 μg/ml (colored green). Nuclear DNA was labelled in blue with DAPI.

  Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069)Lab

  IHC image of Vimentin staining in a section of formalin-fixed paraffin-embedded human normal kidney* performed on a Leica Biosystems BOND® RX instrument. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20mins. The section was then incubated with ab8069, 0.5ug/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. The inset secondary-only control image is taken from an identical assay without primary antibody.

  For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre.

• 

  Flow Cytometry (Intracellular) - Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069)

  Flow cytometry overlay histogram showing wild-type HAP1 (green line) and VIM knockout HAP1 cells stained with ab8069 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized 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 (ab8069) (1x106 in 100 µl at 0.04 µg/ml) for 30 min at 22°C.

  The secondary antibody Goat anti-mouse IgG H&L (Alexa Fluor® 488, pre-adsorbed) (ab150117) was used at 1/2000 for 30 min at 22°C.

  Isotype control antibody was mouse IgG1&kappa (ab170190) used at the same concentration and conditions as the primary antibody (wild-type HAP1 - black line VIM 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 can also be used in HAP1 cells fixed with 4% formaldehyde (10 min) / permeabilized with 0.1% PBS-Triton X-100 for 15 min used under the same conditions.

• 

  Western blot - Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069)

  All lanes : Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069) at 1 µg/ml
  
  Lane 1 : Wild-type HAP1 whole cell lysate
  Lane 2 : VIM (Vimentin) knockout HAP1 whole cell lysate
  Lane 3 : HeLa whole cell lysate
  Lane 4 : MOLT4 whole cell lysate
  
  Lysates/proteins at 20 µg per lane.
  
  Predicted band size: 54 kDa
  
  
  
  

  Lanes 1 - 4: Merged signal (red and green). Green - ab8069 observed at 57 kDa. Red - loading control, ab181602, observed at 37 kDa.
  
  ab8069 was shown to specifically react with Vimentin in wild-type HAP1 cells as signal was lost in VIM (Vimentin) knockout cells. Wild-type and VIM (Vimentin) knockout samples were subjected to SDS-PAGE. Ab8069 and ab181602 (Rabbit anti-GAPDH loading control) were incubated overnight at 4°C at 1 μg/ml and 1/10000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed ab216772 and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed ab216777 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

• 

  Western blot - Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069)Image from Tange S et al., PLoS One. 2014;9(12):e115684. Fig 9(H).; doi: 10.1371/journal.pone.0115684. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

  All lanes : Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069)
  
  Lane 1 : Whole cell lysate of A549 cells
  Lane 2 : Whole cell lysate of A549 cells treated with TGF-beta
  Lane 3 : Whole cell lysate of A549 cells overexpressing JARID2
  Lane 4 : Whole cell lysate of A549 cells overexpressing JARID2 treated with TGF-beta
  
  Predicted band size: 54 kDa
  
  

• 

  Western blot - Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069)

  All lanes : Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069) at 1 µg/ml
  
  Lane 1 : HeLa (Human epithelial cell line from cervix adenocarcinoma) Whole Cell Lysate
  Lane 2 : MOLT4 (Human lymphoblastic leukemia cell line) Whole Cell Lysate
  
  Lysates/proteins at 10 µg per lane.
  
  Secondary
  All lanes : Goat Anti-Mouse IgG H&L (HRP) preadsorbed (ab97040) at 1/5000 dilution
  
  Developed using the ECL technique.
  
  Performed under reducing conditions.
  
  Predicted band size: 54 kDa
  Observed band size: 57 kDawhy is the actual band size different from the predicted?
  
  
  Exposure time: 20 minutes
  

• 

  Immunocytochemistry/ Immunofluorescence - Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069)

  ab8069 staining Vimentin in HeLa cells. 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 overnight at +4°C with ab8069 at 1/100 dilution (shown in green) and ab195889, Mouse monoclonal to alpha Tubulin (Alexa Fluor® 594), at 2µg/ml (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
  Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• 

  Immunocytochemistry/ Immunofluorescence - Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069)Image from Loessner D et al, Biomaterials. 2010 Nov;31(32):8494-506. Epub 2010 Aug 14. doi:10.1016/j.biomaterials.2010.07.064

  ab8069 staining Vimentin in human epithelial ovarian serous adenocarcinoma cell line SKOV-3 by Immunocytochemistry/ Immunofluorescence.
  Samples were fixed with 4% PFA in PBS pH 7.4 and then permeabilised using 0.2% saponin for 30 minutes. A blocking step was performed using 1% BSA/PBS for 1 hour. Samples were then incubated with ab8069 at a 1/200 dilution in 1% BSA/PBS for 1 hour. The secondary antibody was a goat anti-mouse Alexa 488 (green) diluted 1/1000, 1% BSA/PBS for 1 hour. Samples were then incubated with phalloidin (red for actin staining) in 1% BSA/PBS for 45 minutes and counterstained with DAPI (blue for nuclei staining) in PBS for 45 minutes.

• 

  Immunocytochemistry/ Immunofluorescence - Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069)This image is courtesy of an anonymous Abreview

  ab8069 staining Vimentin in rat glial cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with formaldehyde and blocked with 1% BSA for 10 minutes at 25°C. Samples were incubated with primary antibody (1/200) for 10 hours at 4°C. An Alexa Fluor® 555-conjugated anti-mouse IgG polyclonal (1/300) was used as the secondary antibody.

  See Abreview

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069)This image is courtesy of an anonymous Abreview

  ab8069 staining Vimentin in Human oral cavity tissue sections by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections).
  
  Tissue was fixed with paraformaldehyde and blocked with ab64226 Protein Block for 5 minutes at 25°C; antigen retrieval was by heat mediation in pH 6 buffer . Samples were incubated with primary antibody (1/1000 in 10% NGS) for 16 hours at 4°C. An undiluted biotinylated goat anti-rabbit polyclonal IgG was used as the secondary antibody.

  See Abreview

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069)

  IHC image showing no staining when ab8069 was used on mouse kidney formalin fixed paraffin embedded tissue, using MOM detection kit, ab127055. The section was pre-treated using pressure cooker heat mediated antigen retrieval with sodium citrate buffer (pH6) for 30 mins. The section was incubated with ab8069 at 5 µg/ml for 15 mins at room temperature. DAB was used as the chromogen. The section was counterstained with haematoxylin and mounted with DPX.

  For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

• 

  Flow Cytometry (Intracellular) - Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069)

  Overlay histogram showing SV40LT-SMC cells stained with ab8069 (red line). The cells were fixed with 4% formaldehyde (10 min) and then permeabilized with 0.1% PBS-Triton X-100 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 (ab8069, 0.1μg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was Alexa Fluor® 488 goat anti-mouse IgG (H&L) (ab150113) at 1/2000 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 0.1μg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control.

  Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.

  This antibody gave a positive signal in SV40LT-SMC cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Triton X-100 for 20 min used under the same conditions.

• 

  Flow Cytometry (Intracellular) - Anti-Vimentin antibody [V9] - Cytoskeleton Marker (ab8069)

  Overlay histogram showing MDA-MB-231 cells stained with ab8069 (red line). The cells were fixed with 4% formaldehyde (10 min) and then permeabilized with 0.1% PBS-Triton X-100 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 (ab8069, 1μg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was Alexa Fluor® 488 goat anti-mouse IgG (H&L) (ab150113) at 1/2000 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 1μg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control.

  Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.

  This antibody gave a positive signal in MDA-MB-231 cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Triton X-100 for 20 min used under the same conditions.