概述

  • 产品名称Anti-Hsp70抗体[3A3]
    参阅全部 Hsp70 一抗
  • 描述
    小鼠单克隆抗体[3A3] to Hsp70
  • 特异性ab5439 detects several members of the heat shock protein 70 kDa (Hsp 70) gene family including Hsp 70, Hsc 70, p75, and following heat shock, Hsp 72 from yeast, Drosophila, fish, mouse, avian, amphibian and human samples.
  • 经测试应用适用于: Inhibition Assay, ELISA, Flow Cyt, ICC/IF, ICC, IHC-P, WB, IPmore details
  • 种属反应性
    与反应: Mouse, Rat, Chicken, Cow, Human, Pig, Saccharomyces cerevisiae, Drosophila melanogaster, Fish, Non Human Primates, Plants, Amphibians
    预测可用于: Dog, Bird, Cynomolgus Monkey, African Green Monkey, Bos mutus grunniens
  • 免疫原

    Recombinant fragment corresponding to Human Hsp70 aa 504-617 (N terminal).

  • 表位Epitope mapping with a panel of Hsp 70 deletion mutants suggests that the epitope recognized is located between amino acids 504-617 of human Hsp 70, a region that has been shown to be involved in stress-induced nucleolar localization.

性能

  • 形式Liquid
  • 存放说明Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle.
  • 存储溶液Preservative: 0.05% Sodium azide
    Constituent: 99% PBS
  • Concentration information loading...
  • 纯度Ascites
  • Primary antibody说明The Hsp 70 family is a set of highly conserved proteins that are induced by a variety of biological stresses, including heat stress, in every organism in which the proteins have been examined. The human Hsp 70 family members include: Hsp 70, a protein which is strongly inducible in all organisms but which is also constitutively expressed in primate cells; Hsp 72, a 72 kDa protein that is induced exclusively under stress conditions; Hsc 70, or cognate protein, is a 72 kDa, constitutively expressed, protein which is involved in the uncoating of clathrin coated vesicles; GRP78, or BiP, is a glucose regulated 78 kDa protein localized in the endoplasmic reticulum; and p75, or Hsp 75, a 75 kDa protein that is found within the mitochondria.
  • 克隆单克隆
  • 克隆编号3A3
  • 同种型IgG1
  • 研究领域

应用

Our Abpromise guarantee covers the use of ab5439 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

应用 Ab评论 说明
Inhibition Assay Use at an assay dependent concentration.
ELISA Use at an assay dependent concentration.
Flow Cyt 1/100.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.

ICC/IF 1/50 - 1/200.
EMSA Use at an assay dependent concentration.
ICC 1/100. Immunocytochemical staining of Hsp 70 in heat shocked HeLa cells with ab5439 results in cytoplasmic staining.
IHC-P 1/100. Antigen retrieval is not essential but may optimise staining.
WB 1/1000 - 1/5000. Predicted molecular weight: 70-78 kDa.

Representing different members of the HSP70 family. 2-dimensional gel electrophoresis is required to resolve the heat induced form of these proteins from their constitutively expressed counterparts.

IP Use a concentration of 2 µg/ml.

靶标

Anti-Hsp70 antibody [3A3] 图像

  • Immunofluorescent analysis of Heat Shock Protein 70 using Heat Shock Protein 70 Monoclonal antibody (3A3) ab5439 shows staining in MCF-7 cells. Heat Shock Protein 70 staining (green) F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control) or with or an antibody recognizing Heat Shock Protein 70 ab5439 at a dilution of 1/100-1/200 over night at 4 oC washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. Images were taken at 60X magnification.

  • Immunofluorescent analysis of Heat Shock Protein 70 using Heat Shock Protein 70 Monoclonal antibody (3A3) ab5439shows staining in HeLa cells. Heat Shock Protein 70 staining (green) F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control) or with or an antibody recognizing Heat Shock Protein 70 ab5439 at a dilution of 1/100-1/200 over night at 4oC washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. Images were taken at 60X magnification.

  • Immunofluorescent analysis of Heat Shock Protein 70 using Heat Shock Protein 70 Monoclonal antibody (3A3) ab5439 shows staining in NIH-3T3 cells. Heat Shock Protein 70 staining (green) F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control) or with or an antibody recognizing Heat Shock Protein 70 ab5439 at a dilution of 1/100-1/200 over night at 4oC washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. Images were taken at 60X magnification.

  • All lanes : Anti-Hsp70 antibody [3A3] (ab5439) at 1/2500 dilution

    Lane 1 : U2OS cell lysate
    Lane 2 : HeLa cell lysate

    Lysates/proteins at 30 µg per lane.

    Secondary
    HRP-conjugated Goat polyclonal to Mouse at 1/2000 dilution

    Performed under reducing conditions.

    Predicted band size : 70-78 kDa
    Observed band size : 70 kDa (why is the actual band size different from the predicted?)


    Exposure time : 2 minutes

    This image is courtesy of an Anonymous abreview.

    See Abreview

  • Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized human Breast carcinoma tissue. To expose target proteins, heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer, microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1/50 with a mouse monoclonal antibody recognizing Heat Shock Protein 70 (ab5439) or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP, followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.

  • Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized human Testis tissue. To expose target proteins, heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer, microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1/200 with a mouse monoclonal antibody recognizing Heat Shock Protein 70 (ab5439) or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP, followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.

  • Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized human Tonsil tissue. To expose target proteins, heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer, microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1/20 with a mouse monoclonal antibody recognizing Heat Shock Protein 70 (ab5439) or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP, followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.

  • Overlay histogram showing Jurkat cells stained with ab5439 (red line). The cells were fixed with 4% paraformaldehyde (10 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 (ab5439, 1/100 dilution) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in Jurkat cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

  • Immunoprecipitation of Hsp70 was performed on HeLa cells. Antigen:antibody complexes were formed by incubating 500ug whole cell lysate with 2ul of Hsp70 monoclonal antibody (ab5439) overnight on a rocking platform at 4oC. The immune complexes were captured on 50ul Protein Agarose washed extensively and eluted with Buffer. Samples were then resolved on a 4-20% Tris-HCl polyacrylamide gel and transferred to a PVDF membraneand blocked with 5% BSA/TBST for at least 1 hour. The membrane was probed with a Hsp70 monoclonal antibody (ab5439) at a dilution of 1/1000 overnight rotating at 4oC and probed with goat anti-mouse IgG-HRP secondary antibody at a dilution of 1/20,000 for at least 1 hour. Chemiluminescent detection was performed.

Anti-Hsp70 antibody [3A3] (ab5439)参考文献

This product has been referenced in:
  • Inagaki K  et al. Sublethal Photothermal Stimulation with a Micropulse Laser Induces Heat Shock Protein Expression in ARPE-19 Cells. J Ophthalmol 2015:729792 (2015). ICC/IF ; Human . Read more (PubMed: 26697211) »
  • Xu J  et al. Colorectal cancer cells refractory to anti-VEGF treatment are vulnerable to glycolytic blockade due to persistent impairment of mitochondria. Mol Cancer Ther 12:717-24 (2013). Read more (PubMed: 23427299) »

See all 9 Publications for this product

Product Wall

Application Western blot
Sample Human Cell lysate - whole cell (cell lysates in 25% Nupage LSD, 10% DTT, 65% milli)
Gel Running Conditions Reduced Denaturing (12.5% Acrylamide gel)
Loading amount 35 µg
Specification cell lysates in 25% Nupage LSD, 10% DTT, 65% milli
Blocking step Rockland blocking buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 20°C
Username

Abcam user community

Verified customer

提交于 May 27 2015

Application Western blot
Loading amount 20 µg
Gel Running Conditions Reduced Denaturing (8% acrylamide)
Sample Pig Tissue lysate - other (Muscle sarcoplasmic extract)
Specification Muscle sarcoplasmic extract
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C
Username

Dr. Shannon Cruzen

Verified customer

提交于 Mar 27 2014

Thank you for your enquiry.

I would like to reassure you that both these antibodies are covered by our 6 month guarantee for WB and for human samples.

For more details regarding our guarantee, please see the following page from ou...

Read More

Thank you for attending our webinar on Immunoprecipitation last week. We hope you enjoyed it and it was useful for you. We would also like to thank you for sending your question and apologize that we were not able to answer it in the Q&A session. ...

Read More

Thank you for your message and for confirming these details. I am very pleased to hear the customer would like to accept our offer and test these antibodies in hamster. ab41684 Discount code: #### ab38975 Discount code: #### ab6314 ...

Read More

Thank you for your message which has been forwarded to me as Karin is currently away from the office. I have reviewed these three antibodies with respect to discount codes: ab77179 I am sorry I am also unable to check the alignment of the immun...

Read More

Thank you for contacting us. If an antibody has not been tested with a particular species then a testing discount would apply. However, in order to check the likelihood of the antibody being reactive against the target the hamster protein sequence need...

Read More
Application Western blot
Loading amount 30 µg
Gel Running Conditions Reduced Denaturing (12% Tris Glycine)
Sample Human Cell lysate - whole cell (U2OS and HeLa cells)
Specification U2OS and HeLa cells
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Abcam user community

Verified customer

提交于 Jan 18 2011

Thank you for your email. We do not routinely offer free or trial sized samples for testing purposes. Our policy at Abcam is that if an antibody does not work as specified on the datasheet, we will offer a replacement or reimbursement. Ab5439 re...

Read More

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"