Overview

  • Product name
    Anti-Aquaporin 4 antibody [4/18]
    See all Aquaporin 4 primary antibodies
  • Description
    Mouse monoclonal [4/18] to Aquaporin 4
  • Host species
    Mouse
  • Tested applications
    Suitable for: ELISA, IHC-Fr, IHC-P, WB, IHC-FoFrmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Rabbit, Human, Zebrafish
    Predicted to work with: Cow
  • Immunogen

    Synthetic peptide:

    VIDIDRGDEKKGKDSSGE

    , corresponding to amino acids 301-318 of Aquaporin 4

  • Epitope
    Intracellular C-terminal AQP 4 epitope

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Store at -20°C or -80°C. Avoid freeze / thaw cycle.
  • Storage buffer
    Preservative: 0.097% Sodium azide
    Constituents: PBS, 0.1% BSA
  • Concentration information loading...
  • Purity
    Protein A purified
  • Clonality
    Monoclonal
  • Clone number
    4/18
  • Isotype
    IgG3
  • Light chain type
    kappa
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab9512 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
ELISA Use at an assay dependent concentration. PubMed: 11595449
IHC-Fr Use at an assay dependent concentration.
IHC-P 1/100. Perform heat mediated antigen retrieval via the microwave method before commencing with IHC staining protocol.
WB Use at an assay dependent concentration. PubMed: 19831719
IHC-FoFr Use at an assay dependent concentration. PubMed: 22438975

Target

  • Function
    Forms a water-specific channel. Osmoreceptor which regulates body water balance and mediates water flow within the central nervous system.
  • Tissue specificity
    Brain - muscle >> heart, kidney, lung, and trachea.
  • Sequence similarities
    Belongs to the MIP/aquaporin (TC 1.A.8) family.
  • Domain
    Aquaporins contain two tandem repeats each containing three membrane-spanning domains and a pore-forming loop with the signature motif Asn-Pro-Ala (NPA).
  • Post-translational
    modifications
    Phosphorylation by PKC at Ser-180 reduces conductance by 50%. Phosphorylation by PKG at Ser-111 in response to glutamats increases conductance by 40%.
  • Cellular localization
    Membrane.
  • Information by UniProt
  • Database links
  • Alternative names
    • AQP 4 antibody
    • AQP-4 antibody
    • AQP4 antibody
    • AQP4_HUMAN antibody
    • Aquaporin type 4 antibody
    • Aquaporin-4 antibody
    • Aquaporin4 antibody
    • HMIWC 2 antibody
    • HMIWC2 antibody
    • Mercurial insensitive water channel antibody
    • Mercurial-insensitive water channel antibody
    • MGC22454 antibody
    • MIWC antibody
    • WCH 4 antibody
    • WCH4 antibody
    see all

Images

  • Immunohistochemical staining of aquaporin 4 in rat brain.
  • Immunohistochemical analysis of mouse brain tissue, staining Aquaporin 4 with ab9512 at 1/100 dilution. Staining was detected using a biotin-conjugated anti-mouse antibody followed by FITC-conjugated streptavidin.

References

This product has been referenced in:
  • Krause M  et al. Localization, Occurrence, and CSF Changes of SP-G, a New Surface Active Protein with Assumable Immunoregulatory Functions in the CNS. Mol Neurobiol N/A:N/A (2018). Read more (PubMed: 30032421) »
  • Gu C  et al. Simvastatin improves intracerebral hemorrhage through NF-?B-mediated apoptosis via the MyD88/TRIF signaling pathway. Exp Ther Med 15:377-382 (2018). Read more (PubMed: 29375693) »
See all 16 Publications for this product

Customer reviews and Q&As

1-10 of 17 Abreviews or Q&A

Application
Immunohistochemistry (Frozen sections)
Sample
Rabbit Tissue sections (Brain)
Permeabilization
No
Specification
Brain
Blocking step
NDS as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C
Fixative
Paraformaldehyde

Govindaiah Vinukonda

Verified customer

Submitted Jul 09 2018

Abcam has not validated the combination of species/application used in this Abreview.
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Bos taurus Tissue sections (kindey)
Antigen retrieval step
Heat mediated
Permeabilization
Yes - saponin
Specification
kindey
Blocking step
BSA as blocking agent for 30 minute(s) · Concentration: 1% · Temperature: 20°C
Fixative
Paraformaldehyde

Dr. Katarzyna Michałek

Verified customer

Submitted Apr 18 2018

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Dog Tissue sections (intervertebral Disc)
Antigen retrieval step
None
Permeabilization
No
Specification
intervertebral Disc
Blocking step
1% BSA in 75% TBS 35% Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 25% · Temperature: 18°C
Fixative
10% Neutral Buffered Formalin

Abcam user community

Verified customer

Submitted Jul 14 2017

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Rat Tissue sections (Brain)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Ph9 Microwave
Permeabilization
No
Specification
Brain
Blocking step
Serum as blocking agent for 30 minute(s) · Concentration: 1500µg/mL · Temperature: RT°C
Fixative
Paraformaldehyde

Gaia Brezzo

Verified customer

Submitted Sep 06 2016

Answer

Thank you for this information. It is unusual for one to see a band and then subsequently see nothing, which is why I thought you may have been stripping and re-probing. You may improve your result by increasing the concentration of the primary antibody further. Generally the actin band will always be a lot stronger since it is a highly expressed loading control. We can send you a free of charge replacement of the antibody if you wish to try a new lot. Please let me know if this is satisfactory.

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Question
Answer

I am sorry to hear that this antibody is not providing satisfactory results.
You indicated that the secondary antibody used is a rabbit antibody that reacts with rat, however the primary is in mouse so you would need an anti-mouse antibody. Please check this and let me know if you are using a compatible secondary.

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Question

Inquiry: 1. Order details: - Batch number: GR16622-3 - Abcam product code: ab9512 - Antibody storage conditions: +4c 2. Please describe the problem (high background, wrong band size, more bands, no band etc). More bands 3. On what material are you testing the antibody in WB? - Species: rat - What’s cell line or tissue: brain cortex - Cell extract or Nuclear extract: cell extract - Purified protein or Recombinant protein: 3. The lysate - How much protein was loaded: 30ug - What lysis buffer was used: RIPA - What protease inhibitors were used: Roche protease inhibitors cocktail - What loading buffer was used: protein sample buffer - Phosphatase inhibitors: no - Did you heat the samples: 95゚C 10min 4. Electrophoresis/Gel conditions/ Transfer conditions - Reducing or non reducing gel: SDS-page - Reducing agent: 2-mercaptoethanol - Gel percentage : 12% - Transfer conditions: (Type of membrane, Protein transfer verified): semi-dry 5. Blocking conditions - Buffer: PBST - Blocking agent: milk, BSA, serum, what percentage: 5%milk - Incubation time: 1hr - Incubation temperature: room temp 6. Primary Antibody - Species: mouse - Reacts against: rat - At what dilution(s) have you tested this antibody:1:750 - What dilution buffer was used: PBST - Incubation time: overnight - Incubation temperature: 4゚C - What washing steps were done: 3 times per 5 min 7. Secondary Antibody - Species:goat - Reacts against: mouse - At what dilution(s) have you tested this antibody: 1:5000 - Incubation time: 1 hr - Wash steps: 5 times per 5 min - Fluorochrome or enzyme conjugate: HRP - Do you know whether the problems you are experiencing come from the secondary? no 8. Detection method ECl, ECl+, other detection method: ECL 9. Did you apply positive and negative controls along with the samples? Please specify. 10. Optimization attempts - How many times have you tried the Western? 5 - Have you eliminated the possibility that any background bands could be due to the secondary antibody? (Run a “No primary” control): yes - Do you obtain the same results every time e.g. are background bands always in the same place? yes - What steps have you altered? no

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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 would like to offer some suggestions to help optimize the results from ab9512 Anti-Aquaporin 4 antibody [4/18]. I would also appreciate if you can confirm some further details:


1. As this is a multipass membrane protein, which tent to aggregate in the loading buffer when boiled at high temperature due to their hydrophobic nature, I would like to recommend lowering the boiling temperature down to 60C, but keepingthe boiling time of10 min. In addition, use DTT instead of beta-mercaptoethanol as this kind of aggregation is favoured by b- ME.

2. Please be aware that there are two isoforms of this protein, one is 29 and one is 35 kDa.

3.For a better saturation of the membrane, I would recommend a blocking buffer containing 3% BSA for 2 hours at room temperature. BSA can sometimes give better results than milk.Please use the same agent during antibody incubation.

3. In order to increase the specific binding, I would like to recommend decreasing the antibodies concentration down 1:1500.


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.

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Answer

Thank you for contacting us.

We realize that our products can be considered to have premium price point. However, we feel that the product and service which we offer are of the highest possible quality. Under our Abpromise guarantee, ourproducts are guaranteed to work in the manner stated on the datasheetfor a period of six months. If you experience any difficulties using our product and if you have purchased this product within the last 180 days, we are happy to provide scientific support, refund or replacement.

We do offer bulk discounts for orders of 10 units or more, and often run promotions for a number of product types. Please see our website or speak with customer service representative for more information about these.

I hope that this information is helpful. Please let me know if you have any questions or there are other ways that Abcam may help you meet your research goals.

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Answer

Thank you for contacting us. I am glad to hear that this replacement product is performing better than the last product.

Many factors can alter the predicted molecular weight of a protein. These may include post-translational modifications, protein processing, isoforms from alternative splicing, multimer formation, and amino acid charge.

Non-specific binding of an antibody to proteins other than the antigen can sometimes occur. This is usually more common with polyclonal antibodies, but can also occur with monoclonals as well.

An immunizing peptide blocking experiment can be performed to determine which band or staining is specific. As you have mentioned, this product's immunogen is available for use as a blocking peptide. I would recommend using this if you are concerned about the specificity of this product.

In case you would like more information regarding the use of blocking peptides, I have included Abcam's protocol for blocking with the immunizing peptide.

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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Answer

Thank you for your enquiry.

I would like to reassure you that our antibodies have all been tested successfully and would be covered by our guarantee in the applications listed on the datasheets. Therefore, any Aquaporin 4 antibody tested in the species and application you are using should be suitable for your applications.

Also, although we produce many in-house antibodies, we obtain other antibodies from a wide range of sources. Therefore, comparison experiments will not often have been done. I would recommend that when a user is deciding upon a product that they look for those products which have Abreviews and have been used in publications, if possible. When choosing an antibody to be used in Western blot or Flow Cytometry I would also recommend choosing a product which has either a peptide available or the peptide sequence published as the peptide can be used for blocking if necessary.

Please let me know which product your customer would prefer. I look forward to hearing from you. Should you have any further questions please do not hesitate to ask.

Thank you for contacting us.






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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1-10 of 17 Abreviews or Q&A

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