Overview

  • Product name
    Anti-Pericentrin 1/FROUNT antibody
    See all Pericentrin 1/FROUNT primary antibodies
  • Description
    Goat polyclonal to Pericentrin 1/FROUNT
  • Host species
    Goat
  • Tested applications
    Suitable for: ICC/IF, WB, ELISAmore details
  • Species reactivity
    Reacts with: Human
    Predicted to work with: Mouse, Rat
  • Immunogen

    Synthetic peptide corresponding to Human Pericentrin 1/FROUNT aa 644-656 (C terminal).
    Sequence:

    ARAIIREGSLEGS

  • Positive control
    • WB: Hela lysate. ICC/IF: T47D cells.
  • General notes

    Protein previously labeled as Pericentrin 1.

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid freeze / thaw cycles.
  • Storage buffer
    Preservative: 0.02% Sodium Azide
    Constituents: 0.5% BSA, Tris buffered saline. pH 7.3
  • Concentration information loading...
  • Purity
    Immunogen affinity purified
  • Purification notes
    Purified from goat serum by ammonium sulphate precipitation followed by antigen affinity chromatography using the immunizing peptide.
  • Clonality
    Polyclonal
  • Isotype
    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab19044 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
ICC/IF Use at an assay dependent concentration. PubMed: 19841162
WB Use a concentration of 1 - 3 µg/ml. Detects a band of approximately 75 kDa (predicted molecular weight: 75 kDa).
ELISA 1/32000.

Target

  • Function
    Essential component of the nuclear pore complex (NPC) that seems to be required for NPC assembly and maintenance. As part of the NPC Nup107-160 subcomplex plays a role in RNA export and in tethering NUP98/Nup98 and NUP153 to the nucleus. The Nup107-160 complex seems to be required for spindle assembly during mitosis. NUP85 is required for membrane clustering of CCL2-activated CCR2. Seems to be involved in CCR2-mediated chemotaxis of monocytes and may link activated CCR2 to the phosphatidyl-inositol 3-kinase-Rac-lammellipodium protrusion cascade.
  • Sequence similarities
    Belongs to the nucleoporin Nup85 family.
  • Cellular localization
    Nucleus > nuclear pore complex. Chromosome > centromere > kinetochore. Cytoplasm > cytoskeleton > spindle. Cytoplasm. Nucleus membrane. During mitosis, localizes to the kinetochores and spindle poles. Upon CCl2 stimulation translocates from the cytoplasm to the membrane and colocalizes with CCR2 at the front of migrating cells.
  • Information by UniProt
  • Database links
  • Alternative names
    • 85 kDa nucleoporin antibody
    • FLJ12549 antibody
    • FROUNT antibody
    • Frount protein antibody
    • Nuclear pore complex protein nucleoporin 85 kDa antibody
    • nuclear pore complex protein nucleoporin Nup75 antibody
    • Nuclear pore complex protein Nup85 antibody
    • Nucleoporin 85 antibody
    • Nucleoporin 85kDa antibody
    • Nucleoporin nucleoporin 85kDa antibody
    • Nucleoporin Nup75 antibody
    • Nucleoporin NUP85 antibody
    • nucleoporin, 75-KD antibody
    • Nucleoporin85 antibody
    • Nup 75 antibody
    • NUP 85 antibody
    • Nup75 antibody
    • nup85 antibody
    • NUP85_HUMAN antibody
    • PCN antibody
    • PCNT 1 antibody
    • PCNT antibody
    • PCNT, formerly antibody
    • PCNT1 antibody
    • Pericentrin antibody
    • Pericentrin, formerly antibody
    • Pericentrin-1 antibody
    • Pericentrin1 antibody
    see all

Images

  • ab19044 stained T47D cells. The cells were 4% formaldehyde fixed for 10 minutes at room temperature and then incubated in 1%BSA  / 0.3M glycine in 0.1% PBS-Tween for 1hour at room temperature to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab19044 at 1µg/ml) overnight at +4°C. The secondary antibody (pseudo-colored green) was Donkey Anti-Goat IgG H&L (Alexa Fluor® 488) preadsorbed (ab150133) used at a 1/1000 dilution for 1hour at room temperature. Alexa Fluor® 594 WGA was used to label plasma membranes (pseudo-colored red) at a 1/200 dilution for 1hour at room temperature. DAPI was used to stain the cell nuclei (pseudo-colored blue) at a concentration of 1.43µM for 1hour at room temperature.

  • ab19044 (1µg/ml) staining of Hela lysate (35µg protein in RIPA buffer) approx. 75kDa.  Primary incubation was 1 hour. Detected by chemiluminescence.

     

References

This product has been referenced in:
  • Toda E  et al. FROUNT is a common regulator of CCR2 and CCR5 signaling to control directional migration. J Immunol 183:6387-94 (2009). WB, ICC/IF ; Human . Read more (PubMed: 19841162) »
See 1 Publication for this product

Customer reviews and Q&As

1-6 of 6 Abreviews or Q&A

Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (heart)
Specification
heart
Fixative
Paraformaldehyde
Permeabilization
Yes - saponin
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 25°C

Derek Passer

Verified customer

Submitted Aug 13 2012

Question
Answer

DISCOUNT CODE: ***
Expiration date: November 19, 2012

This code will give you 1 free primary antibody before the expiration date. To redeem this offer, please submit an Abreview for ab19044 in IHC-Fr with mouse tissue, and include this code in the “Additional Comments” section so we know the Abreview is for this promotion. Please remember that submission of the Abreview is sufficient for the discount code to become active. For more information on how to submit an Abreview, please visit the site: www.abcam.com/Abreviews.

The code will be active once the Abreview has been submitted and can be redeemed in one of the following ways: 1) Call to place your order and mention the code to our customer service department; 2) Include the code in your fax order; 3) Place your order on the web and enter the promotional code.

Any feedback that you can provide will be greatly appreciated, whether positive or negative. If you have any further questions, please do not hesitate to contact us. We look forward to receiving your Abreview and wish you luck with your research.

The terms and conditions applicable to this offer can be found here: www.abcam.com/collaborationdiscount.

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Question

I am sorry to say that the goat polyclonal anti-Pericentrin 1 antibody ab19044 has indeed not worked on our human samples. We have tried it with three different secondary antibodies. Below I attach a couple of pictures.
This is the second or third centrosome marker from abcam that I try without succeeding. I must say that samples from others have actually worked, so the problem is not our technique. Finally, I would like to mention that the reference given for the goat polyclonal anti-Pericentrin 1 antibody ab19044 in your web page (see below) does not show any picture of this antibody. Also, there is no mention of it in the text nor in the Materials&Methods section.
References for Anti-Pericentrin 1 antibody (ab19044)
This product has been referenced in:

Toda E et al. FROUNT is a common regulator of CCR2 and CCR5 signaling to control directional migration. J Immunol 183:6387-94 (2009). WB, ICC/IF; Human.http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=19841162&dopt=Abstract
Besides that, I have been receiving some emails from your Credit Control Office in which we are asked to pay 10 euros, which account for the different prices of the antibodies we have tried and that didn't work.

I would like to replace the goat polyclonal anti-Pericentrin 1 antibody ab19044 for Anti-GAPDH antibody [mAbcam 9484] - Loading Control (ab9484), which we know ifor sure that works. I do not know if there will be something left to be paid after that last change, but in this regard I also would appreciate that you consider the time invested in trying these antibodies.
Centrine (centrioles) in green. Goat pericentrine (red) does not show any signal.

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Answer

Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products.
I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused.
As you may find in database like SwissProt (ID number Q9BW27, http://www.uniprot.org/uniprot/Q9BW27) or also on the datasheet of ab19044, Pericentrin 1 has several alternative names, including FROUNT. Thus, the publication by Toda and colleagues ('FROUNT is a common regulator of CCR2 and CCR5 signaling to control directional migration.') is indeed a reference describing the use of this antibody in IF with human samples (Fig.5).
I appreciate the time and resources you have spent in the laboratory and understand your concerns. It is regrettable the results have not been successful. As you know, we take product complaints very seriously, and investigate every product that we feel may not be performing correctly. I would like to reassure you that we monitor customer feedback closely on a weekly basis - for all our products - and we are not currently concerned about the general quality of this antibody. I hope you can nevertheless find a suitable antibody and get the experiment working to obtain good results. I wish you the best of luck for your research.
Also, I am very sorry how it worked out with the credit note and the remaining 10 EUR. Please be reassured that the replacements that we issue (for example this present one with ab9484) remain free of charge (so no shipping charges either), even if there is a price difference in the chosen primary antibodies. The credit notes you receive though (for example for the previously not working ab28144) only cover the costs of the original order. Thus, any price difference needs to be pay, may they be as low as 10 EUR. In order to avoid these disappointments in the future, I may suggest to rather accept the free of charge replacement (for example with an alternative product). If you would like to carefully choose the alternatives, I am happy to let you know that we can wait for your decision as long as you like. Just drop us an email and let us know.
As requested, I have issued a free of charge replacement with ab9484 with the order number x.
To check the status of the order please contact our Customer Service team and reference this number.
Please note that this free of charge replacement vial is also covered by our Abpromise guarantee. Should you still be experiencing difficulties, or if you have any further questions, please do not hesitate to let us know.
I hope this experience will not prevent you from purchasing other products from us in the future. Our Scientific Support team is always at your service, should you require further expert advice.

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Answer

Thank you for contacting us and for alerting us to the problem you are experiencing with our mouse monoclonal pericentrin antibody ab28144. We take product complaints very seriously, and investigate every product that we feel may not be performing correctly. We have received no complaints about this product before therefore I am concerned it is again not working - we will certainly follow this up! Indeed, we have a goat pericentrin antibody available, however, I am happy to let you know that we do guarantee it to work ab19044 with human samples in IF, as stated on the datasheet. We just do not have an image available yet, as this antibody has not been tested in-house. If you wish to try it, I would be pleased to arrange a free of charge replacement with this antibody for you. Otherwise, I would of course arrange a refund for you. Thank you for your cooperation. I look forward to hearing from you with details of how you would like to proceed. PS: We are happy to let you know that from now on we offer our Scientific Support also in Spanish language! Thus, if in future you prefer to communicate in Spanish, our colleague Ariana is happy to help either by phone or email.

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Answer

Thank you for your patience. I am sorry to hear that you are experiencing difficulties with this product ab19044 in western blot. Often it is possible to make suggestions that help resolve problems. We will happily offer technical support and in the event that a product is not functioning in the applications cited on the product data sheet (and the problem has been reported within 120 days of purchase), and if it appears that the antibody is at fault, a replacement/credit note/refund will be offered. After looking at the protocols you used, I am surprised that even 40ug of protein, you still did not manage to get any bands. However, it is very hard for me to determine the cause without more details of your WB protocol. I would therefore appreciate if you can please clarify the following items. What animal species was the retinal extract from? Are they known to have sufficient levels for detection? Most customers who use milk as a blocking agent tend to have problems detecting bands. At Abcam, almost all of our products have been tested with 5% BSA. Sometimes, a different blocking buffer might cause cross reaction between the blocking agent and antibodies, therefore causing weak bands or no bands at all. I would recommend trying 5% BSA, if you have not already done so. Also, cross reaction between the antibodies with the diluent could also cause the lack of signals. We would normally use 1% BSA to dilute the primary and secondary antibodies. You can also use PBST only. Can you confirm that the second antibody you used also managed to produce good results with other primary antibodies? In some cases, the problem may be with the secondary antibody not working. Have you tried incubating the primary antibody overnight at 4oC? Incubation at this temperature and for longer duration will ensure proper and sufficient binding to the protein of interest. How long did you keep the antibody at +4°C? Storage at this temperature is only recommended for short term duration (1-2 weeks). Please aliquot and store at -20°C or -80°C for longer storage and avoid repeated freeze / thaw cycles. Did you reduce and denature the sample at 95oC for 10 minutes in buffer containing SDS and mercaptoethanol? This will ensure that the protein is in the correct conformation to run at the correct molecular weight and be detected by the antibody. Lastly, can you confirm the detection method used? At Abcam, we recommend using an ECL+ or super signal kits from Pierce as they are much more sensitive. I am sorry for the string of questions but I hope the above recommendations may already help you. If you have already tried the above suggestions and still experience problems, please do not hesitate to contact me with details of your order (shipping address/purchasing agent information). Also, please advice me on how you would like to proceed with your enquiry, so that I can immediately arrange for a replacement or refund to you.

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Answer

Thank you for your enquiry. I am sorry to hear that you are experiencing difficulties with this product ab19044 in ICC/IF. Often it is possible to make suggestions that help resolve problems. We will happily offer technical support but please bear in mind that this product has not been tested in IHC application and therefore, might not be suitable for use. I have looked at some publications (one listed below) using a clone antibody, and it seems that permeabilization is necessary in order to obtain staining. Please try permeabilizing your samples before incubating with the primary antibody. You can incubate the samples for 10 min with PBS containing 0.25% Triton X-100 (or 100 μM digitonin or 0.5% saponin). Triton X-100 is the most popular detergent for improving the penetration of the antibody. Hansa Patel et al., 2005. Activity and subcellular compartmentalization of peroxisome proliferator-activated receptor are altered by the centrosome-associated protein CAP350. Journal of Cell Science 118, 175-186 Can you confirm that the second antibody you used also managed to produce good results with other primary antibodies? In some cases, the problem may be with the secondary antibody not working properly. If the above suggestions did not help to improve your results, then I am afraid that this product is indeed to not suitable for use in ICC/IF application. In any case, can I encourage you to submit an Abreview via the online product datasheet. We always encourage customers to send their results back to us, whether positive or negative, and we make all product information available to other researchers. We reward each Abreview with Abpoints which can be used for discounts off future purchases and gifts. If you decide to submit an Abreview, please include a note that you have already spoken to me, so that I will be able to quickly publish your Abreview, as I have already tried troubleshooting with you. To find out more about our Abreview system, please see the following URL: https://www.abcam.com/abreviews Should you require any further information or assistance, please do not hesitate to contact me.

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