Overview

  • Product name
  • Description
    Rabbit polyclonal to PC1/3
  • Host species
    Rabbit
  • Tested applications
    Suitable for: IP, IHC-P, WBmore details
  • Species reactivity
    Reacts with: Mouse
    Predicted to work with: Rat, Cow, Human
  • Immunogen

    Synthetic peptide corresponding to Mouse PC1/3 aa 111-127.
    Sequence:

    SVQKDSALDLFNDPMWN


    (Peptide available as ab5011)

  • Positive control
    • WB: mouse pituitary extract IHC-P: FFPE mouse small intestine tissue sections.

Properties

Applications

Our Abpromise guarantee covers the use of ab3532 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
IP Use at an assay dependent concentration.
IHC-P Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
WB Use a concentration of 0.4 µg/ml. Detects a band of approximately 87 kDa (predicted molecular weight: 80 kDa).Can be blocked with PC1/3 peptide (ab5011).

Target

  • Function
    Involved in the processing of hormone and other protein precursors at sites comprised of pairs of basic amino acid residues. Substrates include POMC, renin, enkephalin, dynorphin, somatostatin and insulin.
  • Involvement in disease
    Defects in PCSK1 are the cause of proprotein convertase 1 deficiency (PC1 deficiency) [MIM:600955]. PC1 deficiency is characterized by obesity, hypogonadism, hypoadrenalism, reactive hypoglycemia as well as marked small-intestinal absorptive dysfunction It is due to impaired processing of prohormones.
  • Sequence similarities
    Belongs to the peptidase S8 family. Furin subfamily.
  • Cellular localization
    Cytoplasmic vesicle > secretory vesicle. Localized in the secretion granules.
  • Information by UniProt
  • Database links
  • Alternative names
    • BDP antibody
    • NEC 1 antibody
    • NEC1 antibody
    • NEC1_HUMAN antibody
    • Neuroendocrine convertase 1 antibody
    • PC1 antibody
    • PC3 antibody
    • PCSK1 antibody
    • Prohormone convertase 1 antibody
    • Prohormone convertase 1/3 antibody
    • Prohormone convertase 3 antibody
    • Proprotein convertase 1 antibody
    • SPC3 antibody
    see all

Images

  • IHC image of PC1/3 staining in Mouse small intestine formalin fixed paraffin embedded tissue section, performed on a Leica Bond system using the standard protocol B. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab3532, 1µg/ml, for 15 mins at room temperature. A goat anti-rabbit biotinylated secondary antibody was used to detect the primary, and visualized using an HRP conjugated ABC system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DAB. For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  • Western blot detection of PC1/3 from mouse pituitary extract using ab3532.
  • Anti-PC1/3 antibody (ab3532) at 1 µg/ml + Spinal Cord (Mouse) Tissue Lysate at 10 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 80 kDa
    Observed band size: 87 + 68 kDa
    why is the actual band size different from the predicted?
    Additional bands at: 35 kDa, 60 kDa. We are unsure as to the identity of these extra bands.


    Exposure time: 4 minutes


    PC1/3 contains a number of potential glycosylation sites (SwissProt) which may explain its migration at a higher molecular weight than predicted. The band observed at 68 kDa could potentially be a cleaved form of PC1/3 due to the presence of a 27 amino acid signal peptide and an 83 amino acid propeptide.

References

This product has been referenced in:
  • Irwin N  et al. Chemical cholecystokinin receptor activation protects against obesity-diabetes in high fat fed mice and has sustainable beneficial effects in genetic ob/ob mice. Biochem Pharmacol : (2012). Read more (PubMed: 23085436) »
  • Gagnon J  et al. Expression of PCSK1 (PC1/3), PCSK2 (PC2) and PCSK3 (furin) in mouse small intestine. Regul Pept 152:54-60 (2009). Read more (PubMed: 18706454) »
See all 2 Publications for this product

Customer reviews and Q&As

1-3 of 3 Abreviews or Q&A

Abcam has not validated the combination of species/application used in this Abreview.
Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (mouse brain (hypothalamus))
Permeabilization
No
Specification
mouse brain (hypothalamus)
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: RT°C
Fixative
Paraformaldehyde

Ali Seifinejad

Verified customer

Submitted May 02 2017

Answer

I have received the following information from the scientist who tested ab3532: "The alignment of the C-terminal peptides shows non cross reactivity to mouse is not surprising since there are not 5 contiguous amino acids that are identical in both sequences of mouse and human". I have updated this information on the datasheet and believe therefore that a replacement vial will not give you better results than the original vial. Would you like to have a replacement vial of another antibody? I look forward to hearing from you to arrange a new free order,

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Question

BATCH NUMBER 90293 ORDER NUMBER 147797 DESCRIPTION OF THE PROBLEM no bands SAMPLE STC-1 cells (mouse intestinal endocrine) lysates PRIMARY ANTIBODY abcam rabbit anti-PC1(ab12275) or abcam rabbit anti-PC2(ab3532), 1:2500 or 1:500, 0.01%tween-20/PBS, 1hr RT or overnight 4C plus 1hr RT washing: 2x 1min, 1x 15min, 3x 5 min DETECTION METHOD ECL Plus (Amersham, cat RPN2132) POSITIVE AND NEGATIVE CONTROLS USED loaded STC-1 lysates ANTIBODY STORAGE CONDITIONS antibody dispensed in 2uL aliquots and stored at -20C SAMPLE PREPARATION lysate buffer: complete mini protease inhibitor (Roche, cat #11836153001) AMOUNT OF PROTEIN LOADED tried 10 and 20ug of total protein lysates per lane ELECTROPHORESIS/GEL CONDITIONS 95C at 5 min, DTT, NuPAGE 4-12% Bis-Tris gel (Invitrogen), 150V 1.5 hrs TRANSFER AND BLOCKING CONDITIONS 10% methanol/NuPAGE transfer buffer, 30 V 1.5 hrs, blocking: 5% skim milk/PBS/ 0.01% tween-20 SECONDARY ANTIBODY goat anti-rabbit HRP conjugated (another company), 1:10,000, diluted in 0.01% tween-20/PBS washing: 2x 1min, 1x 15min, 3x 5 min HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 3 HAVE YOU RUN A "NO PRIMARY" CONTROL? No DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes WHAT STEPS HAVE YOU ALTERED? 1) increased primary antibody conc from 1:2500 to 1:500 2) increased incubation time from 1hr RT to overnight at 4c plus 1hr RT 3) increased secondary antibody conc from 1:10,000 to 1:5,000 ADDITIONAL NOTES To test whether any protein was transferred, I probed the same membrane that did not yield any bands with the anti-PC1/PC2 antibodies, with anti-B tubulin and then HRP conjugated secondary. Using this antibody I was able to detect the appropriate bands for this ubiquitous protein.

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Answer

Thank you very much for your protocol details and your order number. I firstly looked at your protocol and think it is very good. I found references for the presence of both PC1 and PC2 in STC-1 cells (Yoon et al, Endocrinol. 1997 for example) and therefore the problem is certainly not due to low amounts of the protein in your samples. However, we have not determined whether ab12275 does cross react with mouse PC2 and I am currently enquiring to find out the exact homology of the immunogen with the mouse protein; I performed a homology search with the last 15 amino acids which contain the immunogen and found 86% homology. I think therefore that it would be important to determine the exact homology so that we can determine if the problem with ab12275 is a species cross reactivity problem. I secondly looked at the shipping records of your order and indeed found a delay; the package was held by regulatory agency on May 24th and I am worried that the package was stored in a hot area. I would like to offer you a replacement vial of ab3532 already and to wait for the cross reactivity information regarding ab12275, as if it is unlikely to cross react with mouse I would recommend to use a different antibody to detect PC2. I appreciate your patience and will e-mail you as soon as I find out the missing information regarding ab12275,

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