Overview

  • Product name
    Anti-Adenosine A1 Receptor antibody
    See all Adenosine A1 Receptor primary antibodies
  • Description
    Rabbit polyclonal to Adenosine A1 Receptor
  • Host species
    Rabbit
  • Specificity
    Detects Adenosine Receptor A1. This antibody does not detect other AR subtypes.
  • Tested applications
    Suitable for: WB, IHC-Pmore details
  • Species reactivity
    Reacts with: Mouse, Rat
    Predicted to work with: Cow, Human
  • Immunogen

    Synthetic peptide corresponding to Rat Adenosine A1 Receptor aa 309-326.
    Sequence:

    CQPKPPIDEDLPEEKAED


    (Peptide available as ab5893)

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle.
  • Storage buffer
    Preservative: 0.05% Sodium azide
    Constituents: 99% PBS, 0.1% BSA
  • Concentration information loading...
  • Purity
    Immunogen affinity purified
  • Primary antibody notes
    Adenosine receptors (ARs) are members of the 7-transmembrane domain G-protein-coupled receptor superfamily. Structural, biochemical and pharmacological analyses of the AR genes and protein has led to the discovery of four distinct AR subtypes (A1, A2a, A2b, A3). Activation of ARs mediates several receptor subtype-specific physiological processes that include cardiac rate, smooth muscle tone, platelet aggregation, inflammation, cell growth and death, and neurotransmission. The A1AR is a glycoprotein that can activate Gi and Go proteins in vitro. In intact cells, agonist occupation of the A1AR has been shown to cause pertussis toxin-sensitive inhibition of adenylyl cyclase activity and, in some systems, a stimulation of phospholipase C resulting in mobilization of intracellular calcium stores. Activation of potassium channels by A1AR has been intensively studied in relation to its dramatic effects on the cardiovascular system. A1AR protein is highly expressed in brain (especially cerebellum, hippocampus, thalamus, and cortex) and spinal cord and in part, modulates neurotransmitter release. In white adipocytes A1AR inhibits lipolysis and stimulates glucose uptake. Other tissues also express A1AR including kidney and testis.
  • Clonality
    Polyclonal
  • Isotype
    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab3460 in the following tested applications.

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

Application Abreviews Notes
WB 1/500. Detects a band of approximately 37 kDa (predicted molecular weight: 37 kDa).Can be blocked with Adenosine A1 Receptor peptide (ab5893).
IHC-P 1/50 - 1/200.

Target

Images

  • ab3460 labelling Adenosine A1 receptor in the cytoplasm of Rat brain tissue (right) compared with a negative control (left) by Immunohistochemistry (formalin/PFA-fixed paraffin embedded sections). To expose target proteins, antigen retrieval method was performed using 10mM sodium citrate (pH 6.0) microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature. Tissue sections were incubated with the primary antibody (1:100 in 3% BSA-PBS) overnight at 4°C. A HRP-conjugated anti-rabbit IgG was used as the secondary antibody, followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

References

This product has been referenced in:
  • Stettler O  et al. Engrailed homeoprotein recruits the adenosine A1 receptor to potentiate ephrin A5 function in retinal growth cones. Development 139:215-24 (2012). Read more (PubMed: 22147955) »
  • Datino T  et al. Mechanisms by which adenosine restores conduction in dormant canine pulmonary veins. Circulation 121:963-72 (2010). WB . Read more (PubMed: 20159830) »
See all 3 Publications for this product

Customer reviews and Q&As

1-4 of 4 Abreviews or Q&A

Answer

Following up my previous message, the immunogen for ab124780, an alternative to ab3460, is also unfortunately unlikely to react with canine Adenosine A1 Receptor. Alignment of the canine receptor amino acid sequence with that of the ab124780 immunogen returns 60% identity. We generally consider this to indicate a low probability of cross-reaction.

Please let me know if we can help in any way.

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Answer

Thank you for contacting us. I think the issue is probably that the homology between the immunogen and the canine receptor is low: 67% identity. We do not have a dog AAR1 to test this.

In general, for multi-pass membrane receptors, we suggest not boiling the sample before loading into the gel , to discourage the aggregation that boiling can induce. Heating at 70C for 10 minutes should be sufficient to denature the sample. (Aggregates do not enter the gel easily). Otherwise, the protocol should be standard, blocking with 5% milk or BSA, and incubation with the antibody diluted to 1/500 in TBST and incubating overnight at 4C. We suggest loading 20-50ug of sample per lane.

Have you considered any of the other AAR1 antibodies we have? None have been tested for reactivity with dog but I will check the immunogen for ab124780 against the dog sequence (http://www.uniprot.org/uniprot/P11616.fasta) and will send the result of the alignment when I have it.

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Answer

Thank you for your reply. If the signal is stronger on frozen section compared to formalin-fixed sections, it indicates that the epitope unmasking may not be proper. We would suggest trying to treat the samples in microwave for different times and to see if the intensity of the signal improves. It is know that certain tissue requires longer retrieval than the other and even if you use a standard well characterized protocol, it may be worth optimizing the technique. As you have suggested, it may well be that the expression level is lower in the lung. One comment on the RT-PCR results - you may need to handle RT-PCR data very carefully. My experience is that high RNA levels do not always correspond with high levels of transcribed protein. If you need anything further or any help then please let us know.

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Answer

Thank you for your enquiry and for your patience. We are very sorry to hear that you are having problem with this antibody. We have searched our database and found that this is a popular selling product and your feedback is the first we have received about it not working. Therefore, at this stage, we would suggest that there is either a problem with the vial you received, or modifications to your protocol are needed to obtain a positive result. A1AR protein is highly expressed in brain (especially cerebellum, hippocampus, thalamus, and cortex) and spinal cord and in part, modulates neurotransmitter release. This antibody was tested in human brain (hippocampus sections) and the pattern is plasma membrane staining. Therefore we would suggest using positive control (i.e. human or rat brain sections) along with the samples. If you use formalin-fixed sections we would suggest antigen retrieval (microwave method), but please make sure that the samples are not over-treated. We would also suggest blocking endogenous peroxidase activity which may give unspecific staining. If you are still having problem with this antibody, please contact us again providing the batch number.

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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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