Overview

  • Product name
    Anti-Pma1 antibody [40B7]
  • Description
    Mouse monoclonal [40B7] to Pma1
  • Host species
    Mouse
  • Specificity
    Reacts with Pma1p based on the size of the band detected on western blots and the immunofluorescence localization pattern.
  • Tested applications
    Suitable for: WB, ICC/IFmore details
  • Species reactivity
    Reacts with: Saccharomyces cerevisiae
  • Immunogen

    Raised against yeast membrane preps, screened by immunofluorescence and blotting.

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 long term. Avoid freeze / thaw cycle.
  • Storage buffer
    Preservative: 0.065% Sodium azide
    Constituent: Tissue culture supernatant
  • Purity
    Tissue culture supernatant
  • Purification notes
    Sterile filtered.
  • Clonality
    Monoclonal
  • Clone number
    40B7
  • Isotype
    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab4645 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 1/10000. Predicted molecular weight: 100 kDa. (cell lysates, ECL detection). For non-ECL western detection methods, 1/1000 - 1/5000. Also detects Pma1p-containing fusion proteins on western blots.
ICC/IF Use at an assay dependent dilution.

Target

  • Relevance
    Pma1p is a plasma membrane ATPase.
  • Database links
    • Alternative names
      • Plasma membrane ATPase 1 antibody
      • Plasma membrane H+ ATPase antibody
      • Pma 1 antibody
      • Pma1p antibody
      • Proton pump 1 antibody
      see all

    Images

    • For Pma1 staining cells were grown to a density of 1 A600/ml, rapidly fixed by direct addition of formaldehyde for 30 minutes, and then harvested and fixed overnight with 4.4% formaldehyde in 0.1 m potassium phosphate, pH 6.5. Before staining, fixed cells were permeabilized with 2% SDS. They were then stained with ab4645 followed by incubation with goat anti-mouse IgG conjugated to Alexa Fluor 488. Stained cells were visualized under fluorescein fluorescence optics on a Zeiss Axioplan 2 fluorescence microscope. Nomarski images (left panels) and indirect immunofluorescence (right panels) are shown for each field.

    References

    This product has been referenced in:
    • Geva Y  et al. Two novel effectors of trafficking and maturation of the yeast plasma membrane H+ -ATPase. Traffic 18:672-682 (2017). WB . Read more (PubMed: 28727280) »
    • Simpkins JA  et al. Disruption of a cystine transporter downregulates expression of genes involved in sulfur regulation and cellular respiration. Biol Open 5:689-97 (2016). WB . Read more (PubMed: 27142334) »
    See all 19 Publications for this product

    Customer reviews and Q&As

    1-9 of 9 Abreviews or Q&A

    Question

    Thank you for the reply. Regarding your questions:

    1) Was the transfer to PVDF verified with a Ponceau stain or another method?

    A: I did not Ponceau stain, however the blot was cut in half and the bottom section (approximately 60kD to the bottom) was probed for Pgk1 as a loading control, which showed that there was abundant protein transferred.

    2) Were the secondary antibodies anti-mouse IgG or anti-mouse IgM?

    A: The secondary's used (Goat Anti-Mouse and Donkey Anti-Mouse) were both IgG's (Heavy and Light Chain). I saw that the Pma1 antibody is an IgM antibody, but my understanding is that it should still react with IgG (H+L) secondary ab's, as it would react with the approriate light chain.

    3) What kind of samples you using and what species are they from?

    A: The samples ran ranged from both vegetative and sporulation protein extracts (extracted using alkaline lysis followed by TCA precipitation) as well as crude extracts from subcellular fractionation (supernatant and pellets from 300g, 14,000g, and 100,000g). All samples were from S. cerevisiae.

    I will certainly try both recommendations you had--increasing the protein loaded as well as antibody concentration. How high would you recommend increasing the antibody? My primary concern is that under the conditions I have used I cannot even see a faint band whatsoever. Would you believe that the inability to detect Pma1 had anything to do with solublization?

    Thank you for Abcam's product guarantee regarding replacement, credit, or refund. I have been very satisfied with the other antibodies we purchased from Abcam, and will discuss these options with my PI.

    Thank you for your speedy response,

    Read More
    Answer

    Thank you very much for your reply.

    The anti-IgG (H&L) secondary will bind to the conserved light chains of the IgM primary. I suggest using the secondary at a higher concentration than you normally would with an IgG primary, but I'd expect this secondary to be fine. You can try increasing the primary to 1:500, and the secondary to 1:5000 to see if this helps.

    To confirm solubility of membrane proteins you could try probing for another membrane marker, if you have an antibody to a different target available. I'm not sure of a better way to confirm this, but if the proteins didn't fully extract then this could be the cause of no signal. We do have one other antibody for an S. cerevisiae membrane protein, ab110326-

    https://www.abcam.com/VDACPorin-antibody-16G9E6BC4-Mitochondrial-Loading-Control-ab110326.html

    Would this be helpful for you? I look forward to hearing from you after discussing with your PI. Please let me know if you have any questions or if there is anything else that we can do for you. Have a nice weekend!

    Read More

    Answer

    Thank you very much for contacting us and letting us know about the trouble with this antibody.

    Thank you also for sending so many details about your protocol. I do have a couple of questions, just so I can further understand the situation:

    1) Was the transfer to PVDF verified with a Ponceau stain or another method?
    2) Were the secondary antibodies anti-mouse IgG or anti-mouse IgM?
    3) What kind of samples you using and what species are they from?

    I would expect the antibody to work under the conditions you've described, but it may be helpful to load more protein and increase the concentration of antibody. I understand that you have already tried some troubleshooting, and this antibody is guaranteed to work in WB with S. cerevisiae samples, so if you would like a replacement, credit, or refund, I'll be happy to set that up for you.

    I look forward to hearing from you. If you have any further questions or need anything else, please let me know and I'll be happy to help.

    Read More

    Answer

    Thank you for contacting Abcam.

    Under our Abpromise, we guarantee this antibody to work in western on yeast samples. As we had not tested in oh human samples, then we could not guarantee that it would work on this species.

    I would be very happy to help you troubleshoot though and see if we can try and get the antibody working. A few protocol variations that may help include:

    1 - Block in BSA, as this would help to reduce background and allow a longer exposure to see the 100kDa bands, if the antibody detects human samples.

    2 - Increase the total amount of protein loaded to 40ug.

    3 - Increase the primary antibody concentration from 1/7000, to 1/1000.

    If the above protocol variations do not prove to be successful, then please let me know and we can try and come up with other solutions.

    Also for your future reference, if you would ever like to use on of our antibodies in an untested application or species, then you could take advantage of our testing discount program:

    https://www.abcam.com/index.html?pageconfig=resource&rid=11998&viapagetrap=collaborationdiscount



    Please let me know if there is anything else I can help you with.

    Read More

    Answer

    Thank you for your enquiry.

    Below is a selection of antibodies that are suitable for detecting the plasma membrane and nuclear envelope. They all work in western blot and I would review the datasheets to confirm which species they are tested against.

    Plasma membrane
    ab4645 Anti-Pma1 antibody [40B7]
    www.abcam.com/4645

    ab7671 Anti-alpha 1 Sodium Potassium ATPase antibody [464.6]
    www.abcam.com/ab7671

    ab16505 Anti-pan Cadherin antibody
    www.abcam.com/ab16505

    ab22744 Anti-pan Cadherin antibody [mAbcam22744]
    www.abcam.com/ab22744

    Nuclear envelope
    ab8980 Anti-Lamin A antibody [133A2]
    www.abcam.com/ab8980

    ab8984 Anti-Lamin A + C antibody [131C3]
    www.abcam.com/ab8984

    ab16048 Anti-Lamin B1 antibody
    www.abcam.com/ab16048

    I hope this is useful. Just let me know if you have further questions.

    Read More

    Answer

    Merci beaucoup pour votre intérêt pour nos anticorps. Nous avons trois anticorps actuellement qui pourront correspondre a vos besoin. Veuillez vérifier leur fiche technique s'il vous plaît. Je n'ai pas trouvé un anticorps testé contre pichia pastoris par contre. Dans quels applications est-ce que vous avez besoin d'utiliser cet anticorps? Anti-PDI antibody [PDI-38H8] - ER Marker (ab30319) Click here (or use the following: https://www.abcam.com/index.html?datasheet=30319). ti-Ribophorin I antibody (ab52672) Click here (or use the following: https://www.abcam.com/index.html?datasheet=52672). Anti-KDEL antibody [10C3] (ab94746) Click here (or use the following: https://www.abcam.com/index.html?datasheet=4746). J'espère que ces informations vous sont utiles et que un des anticorps ci dessus peux correspondre à vos besoins. N'hésitez pas à nous recontacter si vous avez des autres questions.

    Read More
    Abcam guarantees this product to work in the species/application used in this Abreview.
    Application
    Western blot
    Sample
    Saccharomyces cerevisiae Cell lysate - other (Yeast membrane preparation)
    Loading amount
    5 µg
    Specification
    Yeast membrane preparation
    Gel Running Conditions
    Reduced Denaturing (7.5%)
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

    Abcam user community

    Verified customer

    Submitted Aug 07 2009

    Question
    Answer

    Thank you for your enquiry. The isotype of this antibody is IgG3 with kappa light chains. I hope this information helps, please do not hesitate to contact us if you need any more advice or information.

    Read More

    Question
    Answer

    Thank you for your enquiry. All the information we have on species cross reactivity is specified on the datasheet, these are updated as soon as any new information is brought to our attention. As far as we are aware, cross reactivity with Canida albican has not yet been tested for use with ab4645. Should you decide to go ahead and purchase this product, please let us know how you get on and in return we will forward a reward of your choice, typically an Amazon gift voucher. Please let me know if you require any further assistance.

    Read More

    Answer

    The Pma1p protein is very antigenic, and antibodies to it turn up frequently if yeast membrane preps are used as the immunogen. However, what the exact antigenic region is, is not currently known.

    Read More

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

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