Hello. We're improving abcam.com and we'd welcome your feedback.

Hello. We're improving abcam.com and we'd welcome your feedback.

Infomation icon

We haven't added this to the BETA yet

New BETA website

New BETA website

Hello. We're improving abcam.com and we'd welcome your feedback.

Take a look at our BETA site and see what we’ve done so far.

Switch on our new BETA site

Now available

Search and browse selected products

  • A selection of primary antibodies

Purchase these through your usual distributor

In the coming months

  • Additional product types
  • Supporting content
  • Sign in to your account
  • Purchase online

Anti-RhoA 抗体 (ab86297)

Reviews (1)Q&A (6)References (20)
Western blot - Anti-RhoA antibody (ab86297)
  • Western blot - Anti-RhoA antibody (ab86297)
  • Western blot - Anti-RhoA antibody (ab86297)
  • Immunocytochemistry/ Immunofluorescence - Anti-RhoA antibody (ab86297)

Key features and details

  • Rabbit polyclonal to RhoA
  • Suitable for: ICC/IF, WB
  • Reacts with: Mouse, Rat, Human
  • Isotype: IgG

リコンビナント抗体で、ロット間での高い再現性を実現

Product image
Anti-Rho antibody [EP487Y] (ab40673)
  • 異なるロット間での安定した再現性
  • 容易なスケールアップ
  • 評価試験による特異性の確認済み
  • 倫理基準に準拠 - アニマル・フリーの生産

製品の概要

  • 製品名

    Anti-RhoA antibody
    RhoA 一次抗体 製品一覧

  • 製品の詳細

    Rabbit polyclonal to RhoA

  • 由来種

    Rabbit

  • アプリケーション

    適用あり: ICC/IF, WBmore details

  • 種交差性

    交差種: Mouse, Rat, Human
    交差が予測される動物種: Chicken, Cow, Dog, Zebrafish, Orangutan

  • 免疫原

    Synthetic peptide corresponding to Human RhoA aa 150 to the C-terminus conjugated to keyhole limpet haemocyanin.
    (Peptide available as ab101307)

  • ポジティブ･コントロール

    • This antibody gave a positive signal in the following whole cell lysates: HL60; MCF7; MDA-MB-231; HeLa; Human Platelets.

  • 特記事項

    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

製品の特性

アプリケーション

The Abpromise guarantee

Abpromise保証は、 次のテスト済みアプリケーションにおけるab86297の使用に適用されます

アプリケーションノートには、推奨の開始希釈率がありますが、適切な希釈率につきましてはご検討ください。

アプリケーション Abreviews 特記事項
ICC/IF
Use a concentration of 10 µg/ml.
WB
Use a concentration of 1 µg/ml. Detects a band of approximately 20 kDa (predicted molecular weight: 21 kDa).
特記事項
ICC/IF
Use a concentration of 10 µg/ml.
WB
Use a concentration of 1 µg/ml. Detects a band of approximately 20 kDa (predicted molecular weight: 21 kDa).

ターゲット情報

  • 機能

    Regulates a signal transduction pathway linking plasma membrane receptors to the assembly of focal adhesions and actin stress fibers. Serves as a target for the yopT cysteine peptidase from Yersinia pestis, vector of the plague, and Yersinia pseudotuberculosis, which causes gastrointestinal disorders. May be an activator of PLCE1. Activated by ARHGEF2, which promotes the exchange of GDP for GTP.

  • 配列類似性

    Belongs to the small GTPase superfamily. Rho family.

  • ドメイン

    The basic-rich region is essential for yopT recognition and cleavage.

  • 翻訳後修飾

    Substrate for botulinum ADP-ribosyltransferase.
    Cleaved by yopT protease when the cell is infected by some Yersinia pathogens. This removes the lipid attachment, and leads to its displacement from plasma membrane and to subsequent cytoskeleton cleavage.
    AMPylation at Tyr-34 and Thr-37 are mediated by bacterial enzymes in case of infection by H.somnus and V.parahaemolyticus, respectively. AMPylation occurs in the effector region and leads to inactivation of the GTPase activity by preventing the interaction with downstream effectors, thereby inhibiting actin assembly in infected cells. It is unclear whether some human enzyme mediates AMPylation; FICD has such ability in vitro but additional experiments remain to be done to confirm results in vivo.
    Ubiquitinated by the BCR(BACURD1) and BCR(BACURD2) E3 ubiquitin ligase complexes, leading to its degradation by the proteasome, thereby regulating the actin cytoskeleton and cell migration.

  • 細胞内局在

    Cell membrane. Cytoplasm > cytoskeleton.
  • Information by UniProt

  • 参照データベース

    see all

  • 別名

    • Aplysia ras related homolog 12 antibody
    • ARH12 antibody
    • ARHA antibody
    • H 12 antibody
    • H12 antibody
    • Oncogene RHO H12 antibody
    • Ras homolog family member A antibody
    • Ras homolog gene family member A antibody
    • Rho A antibody
    • Rho cDNA clone 12 antibody
    • RHO H12 antibody
    • RHO12 antibody
    • RHOA antibody
    • RHOA_HUMAN antibody
    • RHOH12 antibody
    • Small GTP binding protein Rho A antibody
    • Transforming protein Rho A antibody
    • Transforming protein RhoA antibody
    see all

画像

  • Western blot - Anti-RhoA antibody (ab86297)
    Western blot - Anti-RhoA antibody (ab86297)
    All lanes : Anti-RhoA antibody (ab86297) at 1 µg/ml

    Lane 1 : HL60 (Human promyelocytic leukemia cell line) Whole Cell Lysate
    Lane 2 : MCF7 (Human breast adenocarcinoma cell line) Whole Cell Lysate
    Lane 3 : MDA-MB-231 (Human breast adenocarcinoma cell line) Whole Cell Lysate
    Lane 4 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 5 : Human Platelet (Human adult normal cell line) Whole Cell Lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 21 kDa
    Observed band size: 20 kDa why is the actual band size different from the predicted?


    Exposure time: 90 seconds
  • Western blot - Anti-RhoA antibody (ab86297)
    Western blot - Anti-RhoA antibody (ab86297)
    All lanes : Anti-RhoA antibody (ab86297) at 1 µg/ml

    Lane 1 : Kidney (Mouse) Tissue Lysate
    Lane 2 : Mouse lung normal tissue lysate - total protein (ab29297)
    Lane 3 : Kidney (Rat) Tissue Lysate
    Lane 4 : Lung (Rat) Tissue Lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 21 kDa
    Observed band size: 20 kDa why is the actual band size different from the predicted?


    Exposure time: 2 minutes
  • Western blot - Anti-RhoA antibody (ab86297)
    Western blot - Anti-RhoA antibody (ab86297)
  • Immunocytochemistry/ Immunofluorescence - Anti-RhoA antibody (ab86297)
    Immunocytochemistry/ Immunofluorescence - Anti-RhoA antibody (ab86297)
    ICC/IF image of ab86297 stained MCF7 cells. The cells were 4% PFA fixed (10 min) and 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 (ab86297, 10µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899 Dylight 488 goat anti-rabbit IgG (H+L) 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.

参考文献 (20)

ab86297 を使用した論文を発表された方は、こちらまでお知らせください。データシートに掲載させていただきます。

ab86297 は 20 報の論文で使用されています。

  • Yiu TW  et al. Transglutaminase 2 Facilitates Murine Wound Healing in a Strain-Dependent Manner. Int J Mol Sci 24:N/A (2023). PubMed: 37511238
  • Li X  et al. Erianin Controls Collagen-Mediated Retinal Angiogenesis via the RhoA/ROCK1 Signaling Pathway Induced by the alpha2/beta1 Integrin-Collagen Interaction. Invest Ophthalmol Vis Sci 63:27 (2022). PubMed: 35060996
  • Cui Q  et al. Leptin Promotes Angiogenesis via Pericyte STAT3 Pathway upon Intracerebral Hemorrhage. Cells 11:N/A (2022). PubMed: 36078162
  • Wang X  et al. Hypoxia-induced miR-182-5p regulates vascular smooth muscle cell phenotypic switch by targeting RGS5. Cell Biol Int 46:1864-1875 (2022). PubMed: 35946384
  • Tian Y  et al. LncRNA LINC00974 Downregulates miR-122 to Upregulate RhoA in Oral Squamous Cell Carcinoma. Cancer Biother Radiopharm 36:18-22 (2021). PubMed: 31702382
View all Publications for this product

レビューと Q&A

Show All レビュー Q&A
レビューを送る 質問を送る

1-7 of 7 Abreviews or Q&A

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-RhoA antibody

 Excellent
Abreviews
abreview image
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Dog Tissue sections (Uterus)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: citrate buffer pH=6.0
Permeabilization
No
Specification
Uterus
Blocking step
Serum as blocking agent for 20 minute(s) · Concentration: 10% · Temperature: 20°C
Fixative
Formaldehyde
Read More
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Abcam user community

Verified customer

投稿 Oct 14 2016

Question

Thanks for your reply. I did think the size difference is probably due to interactions or modifications. The modifications I am aware of at the moment I expect to be present in the membrane fraction rather than the cytosolic fraction but I will investigate the literature for clues. In the whole cell fraction I only get the 30 kDa band but if the size difference is due to differential properties of membrane and cytosolic RhoA the lack of the smaller band could be explained by RhoA being more dilute in the whole cell fraction than in the membrane. I get my whole cell fraction as part of the fractionation protocol in which I unfortunately can’t use RIPA buffer but I have already added a step to my protocol that will hopefully improve solubilisation if it is the problem. And if this doesn’t help I will try heating with and without SDS and DTT.

Thanks for your help.

Regards,

Read More
Answer

Thank you for your reply.

I hope the use of SDS and DTT will resolve the problem!

Please let us know also if there is anything else we could help you with. We do have for example different cell fractionation kits, and if the SDS and DTT would not clarify the band pattern, we could try sucha kit to see whether the same bands are observed in the different fractions (I problably could make a good testing offer on these if you would then provide a feedback as well).

I wish you all the best for your project and please do not hesitate to contact us again for any questions in relation to our products.

Read More

Question

Sorry it’s taken me such a long time to get back to you. I did manage to get the RhoA antibody working though something slightly unexpected happened and I’m hoping you might be able to help me with this. I am doing subcellular fractionation and depending on the fraction the bands were different sizes; in the nuclear and membrane fractions the bands were about the expected size (or maybe slightly smaller) but in the whole cell and cytosolic fractions the bands were about 30 kDa. The only difference I can think of is the presence of detergent; the nuclear and membrane fractions both have it and not the cytosolic, I have added detergent to the whole cell fraction as well though differently from the nuclear and membrane fractions. Any more info on this would be useful.



Regards,

Read More
Answer

Thank you very much for your email. I am very happy to hear that the RhoA antibody is working now.

The observedband in the cytoplasm is indeed unexpected. I would think this would correspond to a complex in the cytoplasm or another modification.I am however not an expert in RhoA biology and would rely ona more thorough literature research for finding what interaction partner it could be.

In order to analyse howeverfroma technical point of viewthis observed protein, I would recommend to systematically heat with and without SDS and beta Mercaptoethanol (or DTT) the fractions, and see whether the bands are still present. Do you observe the 20kD band also in the whole cell fraction?

What type of buffer have you used to make the cell lysis? I can recommend also to try the RIPA buffer for the whole cell lysate. RIPA buffer is a very good buffer to solubilize the proteins. (https://www.abcam.com/index.html?pageconfig=resource&rid=11379#A1)

Please let me know if this makes sense. I hope this will help to clarify the results. Please keep me updated and do let me know if you would have any doubt about the quality of the antibody.

I am looking forward to hear back from you.

Read More

Question

What is the homology between the immunogen and the mouse RhoB and RhoC protein sequences?

Read More
Answer

Thank you for your call a couple weeks ago, and I sincerely apologize for the delay in getting back to you.
As we discussed, the immunogen of ab86297 is 83% identical to RhoB and 73% to RhoC, so it is possible that this antibody will cross-react. I have unfortunately been unable to obtain any further information about the immunogen of ab68826.

I am sorry that I can not be more help in this situation, but please let me know if there is anything else that I might be able to do for you.

Read More

Question

Thanks for your reply. Yes the samples have been reducedand denaturedand I know there is protein there asb-actin comes up clearly. Also I have used protease inhibitors. I have used the secondary antibody withanotherrabbit primary and that works fine.Ihaven't tried BSA for blocking yet but will do that as soon as possible.

Thanks,

Read More
Answer

Thank you for your email.

I look forward hearing back from you with the results of the BSA blocking.

Have a great day!

Read More

Question

Inquiry: I am not able to get signal from this antibody. I have been testing it in rat liver and have used 20-40 ug of protein. I generally use 4-12% gel as this should be fine for the protein size though I have tried a higher percentage gel as well. Transfers I usually run at 75V for 1.5 hours. I block for 1 hour in room temperature in 5% milk. Primary antibody incubation I do overnight at 4C and I have tried 1:1000 and 1:750 dilutions in milk. Secondary antibody I use at 1:100 000 in RT for 1.5 hours which I know works with another antibody. Washes I do 3 times 7-10 minutes.

Read More
Answer

Thank you for taking time to contact us with this information. I am sorry to hear that this antibody is not providing satisfactory results.

The details provided will enable us to investigate this case and will provide us with vital information for monitoring product quality.

Having reviewed this case, I would like to offer some suggestions to help optimize the results from ab86297. I would also appreciate if you can confirm some further details:

1.) Can you please confirm the samples have been reduced and denatured? Reducing with DTT or beta Mercaptoethanol and denaturing the samples with SDS and heating 5 to 10 minutes at 95°C will help the proteins to enter the gel and resolvenicely.

2.) I can strongly recommend to use BSA as a blocking buffer. Indeed, the laboratory has tested this antibody using BSAfor blocking. Changing the blocking buffer can significantly improve the results. As an illustration, please see also the Western blot image on the datasheet of ab9385 https://www.abcam.com/HRP-GAPDH-antibody-Loading-Control-ab9385.html (or use the following: https://www.abcam.com/HRP-GAPDH-antibody-Loading-Control-ab9385.html).

3.) Can you also confirm, whether the other reagents can be exluded to be at the origin of the problem? - Has the secondary antibody already provided good results with other primary rabbit antibodies?

4.) In order to exclude protein degradation as possible origin of the problem, can you please confirm whether proteases inhibitors have been added to the lysate and whetherother proteins been detected in the same lysates?

I agree also that liver cells should have a high level of RhoA. Should the suggestions not improve the results, please do let me know.

In the event that a product is not functioning in the species and applications cited on the product datasheet (and the problem has been reported within 6 months of purchase), we would be pleased to provide a free of charge replacement, credit note, or refund.

I hope this information is helpful, and I thank you for your cooperation.

Read More

Question

Will this antibody also react with RhoB or RhoC?

Read More
Answer

Thank you for contacting Abcam regarding ab86297. We have not tested this antibody for cross reactivity with RhoB or RhoC.  By blasting the immunogen sequence for this Ab, it shows there may be some cross reactivity with RhoC. I hope this information is helpful.  Please do not hesitate to contact us if you have any additional questions.

Read More

Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
For licensing inquiries, please contact partnerships@abcam.com