製品の概要

  • 製品名
    Anti-beta 1 Adrenergic Receptor antibody
    beta 1 Adrenergic Receptor 一次抗体 製品一覧
  • 製品の詳細
    Rabbit polyclonal to beta 1 Adrenergic Receptor
  • 由来種
    Rabbit
  • アプリケーション
    適用あり: WB, ICC/IF, IP, IHC-FoFr, ICC, IHC-P, Flow Cytmore details
  • 種交差性
    交差種: Mouse, Rat, Cat, Human, Non human primates, African green monkey, Prairie vole
    交差が予測される動物種: Rhesus monkey
  • 免疫原

    Synthetic peptide corresponding to Mouse beta 1 Adrenergic Receptor aa 394-408.
    Sequence:

    HGDRPRASGCLARAG

製品の特性

  • 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. Avoid freeze / thaw cycle.
  • バッファー
    Preservative: 0.05% Sodium azide
    Constituents: 99% PBS, 0.1% BSA
  • 精製度
    Whole antiserum
  • 一次抗体 備考
    Adrenergic receptors (ARs) are members of the 7-transmembrane domain G-protein-coupled receptor superfamily that bind the endogenous catecholamines epinephrine and norepinephrine. Pharmacological, structural, and molecular cloning data indicate significant heterogeneity within this receptor family. Nine receptor subtypes have been identified thus far including three alpha-1 AR subtypes (1A/D, 1B, and 1C), three alpha-2 ARs (2A, 2B, and 2C), and three beta AR subtypes (1, 2, and 3). ARs participate in either the onset or maintenance of several disease states including hypertension, cardiac dysfunction (congestive heart failure, ischemia, arrhythmias), diabetes, glaucoma, depression, and impotence. BARs participate in diverse processes including development, behavior, cardiac function, smooth muscle tone, and metabolism. In gene-knockout experiments, the majority of mice that lack the B1AR gene die prenatally and those that do survive until adulthood display abnormal cardiac function. Other studies have shown that the direct regulation of cardiac B1AR density by thyroid hormones occurs at the transcriptional level and is modulated by the catecholamine sensitive-adenylyl cyclase system. Evidence for the effects of cAMP on B1AR regulation has come from a study of members of the cAMP response element (CRE) modulator (CREM) family of transcription factors in transformed cell lines.
  • ポリ/モノ
    ポリクローナル
  • アイソタイプ
    IgG
  • 研究分野

アプリケーション

Our Abpromise guarantee covers the use of ab3442 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
WB 1/1000. Detects a band of approximately 50.5 kDa (predicted molecular weight: 50 kDa).
ICC/IF 1/100. Immunofluorescence staining of B1AR in mouse kidney distal tubule yields a pattern consistent with plasma membrane staining.
IP Use at an assay dependent concentration.
IHC-FoFr Use at an assay dependent concentration.
ICC 1/5000.
IHC-P 1/200.
Flow Cyt Use at an assay dependent concentration.

ab171870 - Rabbit polyclonal IgG, is suitable for use as an isotype control with this antibody.

ターゲット情報

  • 機能
    Beta-adrenergic receptors mediate the catecholamine-induced activation of adenylate cyclase through the action of G proteins. This receptor binds epinephrine and norepinephrine with approximately equal affinity.
  • 配列類似性
    Belongs to the G-protein coupled receptor 1 family. Adrenergic receptor subfamily. ADRB1 sub-subfamily.
  • ドメイン
    The PDZ domain-binding motif mediates competitive interactions with GOPC, MAGI3 and DLG4 and plays a role in subcellular location of the receptor.
  • 翻訳後修飾
    Homologous desensitization of the receptor is mediated by its phosphorylation by beta-adrenergic receptor kinase.
  • 細胞内局在
    Cell membrane. Localized at the plasma membrane. Found in the Golgi upon GOPC overexpression.
  • Information by UniProt
  • 参照データベース
  • 別名
    • ADR B1 antibody
    • ADRB 1 antibody
    • ADRB1 antibody
    • ADRB1_HUMAN antibody
    • ADRB1R antibody
    • Adrenergic beta 1 receptor antibody
    • Adrenoceptor beta 1 antibody
    • B1AR antibody
    • Beta 1 adrenoceptor antibody
    • Beta 1 adrenoreceptor antibody
    • Beta-1 adrenergic receptor antibody
    • Beta-1 adrenoceptor antibody
    • Beta-1 adrenoreceptor antibody
    • BETA1AR antibody
    • RHR antibody
    see all

画像

  • All lanes : Anti-beta 1 Adrenergic Receptor antibody (ab3442) at 1/1000 dilution

    All lanes :

    Lysates/proteins at 25 µg per lane.

    Secondary
    All lanes : HRP-conjugated Goat anti-Rabbit at 1/20000 dilution

    Predicted band size: 50 kDa
    Observed band size: 50 kDa
    Additional bands at: 30 kDa. We are unsure as to the identity of these extra bands.



    Western blot analysis of ADR-Beta-1 was performed by loading 25ug of various whole cell lysates onto a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a PVDF membrane and blocked with 5% Milk/TBST for at least 1 hour. Membranes were probed with ab3442 at a dilution of 1/1000 overnight at 4°C on a rocking platform. Membranes were washed in TBS-0.1%Tween 20 and probed with a goat anti-rabbit-HRP secondary antibody at a dilution of 1/20,000 for at least one hour. Membranes were washed and chemiluminescent detection performed.
  • ab3442 at 1/100 dilution staining beta 1 Adrenergic Receptor in untreated HeLa cells by Immunocytochemistry/ Immunofluorescence. Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 15 minutes at room temperature. Cells were then blocked with 5% normal goat serum for 15 minutes at room temperature. Cells were probed with a rabbit polyclonal antibody recognizing ADR-Beta-1, at a dilution of 1/100 for at least 1 hour at room temperature. Cells were washed with PBS and incubated with DyLight 488-conjugated goat-anti-rabbit secondary antibody at a dilution of 1/400 for 30 minutes at room temperature. Nuclei were stained with Hoechst 33342 dye.
  • ab3442 staining the beta 1 Adrenergic Receptor in mouse bone marrow cells by Flow Cytometry. Cells were fixed with paraformaldehyde and permeabilized with 0.5% BSA in 2mM EDTA/PBS. The sample was incubated with the primary antibody (1/100) for 20 minutes at 4°C. An Alexa Fluor® 488-conjugated goat anti-rabbit IgG polyclonal (1/1000) was used as the secondary antibody.
    Gating Strategy: Blood cells.

    See Abreview

  • Immunolocalization of B1AR in mouse kidney distal tubule using ab3442.

参考文献

This product has been referenced in:
  • Sun F  et al. Stage-dependent changes of ß2-adrenergic receptor signaling in right ventricular remodeling in monocrotaline-induced pulmonary arterial hypertension. Int J Mol Med 41:2493-2504 (2018). Read more (PubMed: 29393391) »
  • Pan LY  et al. Mechanism of Yanghe Pingchuan granules treatment for airway remodeling in asthma. Drug Des Devel Ther 12:1941-1951 (2018). Read more (PubMed: 29983548) »
See all 16 Publications for this product

レビューと Q&A

1-8 of 8 Abreviews or Q&A

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Blocking step
Serum as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 20°C
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Citrate buffer 10mM, pH=6
Sample
Human Tissue sections (Adipose Tissue)
Specification
Adipose Tissue
Permeabilization
Yes - with ethanol during rehydration of sections
Fixative
Formaldehyde

Abcam user community

Verified customer

投稿 Sep 16 2014

Application
Flow Cytometry
Fixation
Paraformaldehyde
Permeabilization
Yes - 0.1%spaonin/PBS
Sample
Mouse Cell (bone marrow)
Specification
bone marrow
Gating Strategy
blood cells
Preparation
Cell harvesting/tissue preparation method: obtained from murine femur frushed with RPMI
Sample buffer: 0.5% BSA/2mM EDTA/PBS

Abcam user community

Verified customer

投稿 Jun 23 2014

Answer

Thank you for bringing this to our attention. The two bands you are seeing might represent unphosphorylated (lower band) and fully phosphorylated (upper band) receptor but that is unlikely if the difference in sizes is 5 kDa and you are seeing just two bands. There are at least three known phosphorylation sites. There is also the possibility of glycosylation which would cause a portion of the protein to run more slowly than the unmodified portion.

We have not tested this antibody against mouse brain samples, only the cell lysates listed in the caption of the western blot image on the datasheet. The result shows primarily a single band at the 50 kDa marker, and a very light band at 55 for two or three cell lysates. Other antibodies raised against the receptor, for instance ab85037, tested against mouse spinal cord, show just a single band at about 55 kDa. I suspect that one of the bands you see is unrelated to the receptor. We can send a replacement, for instance ab85307 but first I would like to see if there is something I can suggest to improve your ab3442 result.

Can you please send a file of your blot image, and a few details of your protocol? In particular, how was the sample prepared? Did the prep include protease inhibitors?

We have another report of a band appearing at 57 kDa only when the blot is blocked with BSA. When blocked with 5% milk, the band is not present. How did you block the membrane?

Finally, what was the concentration or dilution of the antibody? Have you stained blots of these samples with any other antibodies, and were the results as expected? I look forward to your reply.

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry
Sample
Prairie Vole Cell (brain)
Specification
brain
Fixative
Paraformaldehyde
Permeabilization
No
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 20°C

Abcam user community

Verified customer

投稿 Oct 02 2012

Question


I have used the antibody against b1-adrenergic receptor ab3442 in western blot on human fibroblast cell lysates and obtained a quite unspecific pattern with multiple bands but no major band at the expected molecular weight (50kDa).

I have tried different things: laemmli or RIPA extraction buffers, loading different quantities of protein (10-20ug), blocking in either BSA or milk.

Interestingly, the blocking test gave very different patterns (see attached file):

Milk wasn’t good, showing two major bands at 36 and 38kDa, and multiple weak bands above.

BSA blocking gave, to my mind, the closest result to the expected MW, with a strong band at 57kDa. There was also an even strongest band around 35kDa, (that I noticed is visible on the picture published in the antibody datasheet, with no additional remark about it. Is that a known additional form due to some post-translational modification?)

I got similar multi-band patterns with human lung and dermal fibroblasts, and human endothelial cells.

I have tried to stick to a similar protocol to what is given in the datasheet, but I would be interested to know the exact conditions this antibody has been tested in, especially which lysis buffer was used, and would welcome any advice to improve the protocol.

We have been using a different antibody in the past (ab77189), but the signal was very weak so I wanted to give the ab3442 a try, see if it would be better. Obsviously it’s not… One interesting thing to note is that in ab77189 datasheet, the observed band is 55-60kDa (while it’s still expected at 51kDa). Would the band I detect around 57kDa be the right one?



Many thanks

Kind regards

Read More
Answer

Thank you very much for your inquiry. I apologize for the long delay of my answer.

Firstly, thank you very much for having provided such a nice summary, including the data and the protocol steps. This makes it easier for us, to understand the problem.

I have looked at the data for ab3442 and would like to raise the following points:

1.) When looking at the data you provided, the first thing which comes to my mind is, whether these are not degradation products. Indeed, multitransmembrane protein are very sensitive to heat and can break. Have you heated the samples before you have loaded them on the gel? If yes, I can recommend to test also unheated samples and samples heated just for 5 or 10 minutes at 70°C.

2.) Have the samples been reduced as well?

3.) In order to avoid degradation by proteases as well, have you held the samples always on ice and added protease inhibitors to the RIPA buffer?

4.) I actually can confirm that thelysis buffer you are using (RIPA) should be suitable for membrane proteins.

5.) To my knowledge there are not isoforms described yet for this protein.

6.) The band at 57kD absolutely could be the right band. Depending on the buffer system and the molecular weight marker, there are always slight variation of the apparent molecular weight. Additionally, the Beta-1 adrenergic receptor can also be glycosilated, which increases its apparent molecular weight. http://www.uniprot.org/uniprot/P08588

If with these recommendations we do not resolve the problem, we will replace this antibody for you according to our guarantee. I am however optimistic, that not heating the samples could improve significantly the results. Please let me know how you are getting on and if you have any questions or concerns. I look forward to hear back from you.

Read More

Answer

Thank you for taking time to complete our questionnaire and for contacting us. I am sorry to hear 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. I appreciate the time you have spent in the laboratory and understand your concerns. It is regrettable the results have not been successful.

Having reviewed the protocol details, I believe this product should have given satisfactory results. It appears that you may have received a faulty vial.

I apologize for the inconvenience and am pleased to offer you a free of charge replacement, credit note, or refund in compensation. Unfortunately, we only have the same lot of ab3442 in stock at present to offer. Alternatively I can offer the antibody ab85037. This has shown to work well with mouse spinal chord tissue as well as ICC staining of SHSY5Y cells as displayed on the datasheet of this product.

I look forward to hearing from you with details of how you would like to proceed.

Read More

Question
Answer

Thank you for taking time to complete our questionnaire. 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 wouldappreciate it if you could confirm some further details:

1. Has the secondary antibody and Ecl reagent been used successfully in other blots?

2. Has a loading control been performed with the samples?

3. How long was the membrane exposed for?

I would like to offer some suggestions to help optimize the results from ab3442.

1. When testing our antibodies, our lab uses 5% BSA as a blocking reagent, so I recommend switching to this instead of milk. For unknown reasons some antibodies give stronger, more specific signals on blots blocked with BSA instead of milk, so doing this may improve the results you are seeing, and reduce the non-specific bands. An example of the above is the western blot obtained with the Abcam GAPDH antibody ab9385 : https://www.abcam.com/gapdh-antibody-hrp-loading-control-ab9385.html#GAPDH-Primary-antibodies-ab9385-3.jpg (or use the following: https://www.abcam.com/ab9385).BSA should then also be used in the dilutionbuffer of the antibodies instead of milk.

2. I would suggest reducing the amount of protein loaded onto the gel. 20 ug of protein should be sufficient. I would also suggest heating the sample to 70°C for 10 minutes instead of boiling the sample as the target is a membrane bound protein which tend to form aggregated if boiled.

3. I would also strongly suggest performing a "no primary" control in order to deduce if this non-specificity is due to the secondary antibody or the blocking agent.

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

Answer

Thank you very much for your interest in our adrenergic Receptor antibodies. These are the alternatives we could recommend: Anti-beta 1 Adrenergic Receptor antibody (replacement for ab3546): The rabbit polyclonal ab3442 which is guaranteed to work with a number of species including mouse and rat in Western blot and Immunocytochemistry/Immunofluorescence (ICC/IF). To our knowledge, ab3442 has not been tested in Immunhistochemistry on frozen sections (IHC-Fr). Therefore, I can offer a discount off a future purchase if you buy it now, test it in IHC-Fr and submit feedback to us in the form of an Abreview. It doesn’t matter whether the Abreview is positive or negative, we would just really like to receive your feedback. The discount would be to the value of 1 free primary antibody. Please contact us again for more detailed information if you are interested. Click here (or use the following: https://www.abcam.com/index.html?datasheet=3442). Anti-alpha 1d Adrenergic Receptor antibody (replacement for ab12923): The rabbit polyclonal ab84402 which is guaranteed to work with a number of species including human in Western blot and ELISA. This antibody has not been tested in Immunhistochemistry on paraffin-embedded sections (IHC-P). Again, if you would like to test this, you are eligible for our testing discount program if the antibody has not yet been purchased. Click here (or use the following: https://www.abcam.com/index.html?datasheet=84402). Anti-alpha 1c Adrenergic Receptor antibody (replacement for ab12935): The rabbit polyclonal ab96783 or the mouse monoclonal ab87990 which are guaranteed to work with a number of species including human in Western blot and ELISA. These antibodies have not been tested in Immunhistochemistry on paraffin-embedded sections (IHC-P) and here you are eligible for our testing discount program. Click here (or use the following: https://www.abcam.com/index.html?datasheet=96783). Click here (or use the following: https://www.abcam.com/index.html?datasheet=87990). The terms and conditions applicable to these offers can be found here: www.abcam.com/collaborationdiscount. I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

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

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