Overview

  • Product name
  • Description
    Goat polyclonal to PRPF31
  • Host species
    Goat
  • Tested applications
    Suitable for: WBmore details
  • Species reactivity
    Reacts with: Human
    Predicted to work with: Dog
  • Immunogen

    Synthetic peptide: LKVKGEKSGLMST, corresponding to C terminal amino acids 487-499 of Human PRPF31.

  • Positive control
    • A431 lysate.
  • General notes
    Principal Names - PRPF31; PRP31 pre-mRNA processing factor 31 homolog (yeast); RP11; PRP31; NY-BR-99; DKFZp566J153; pre-mRNA processing factor 31 homolog (yeast). Official Gene Symbol - PRPF31. GenBank Accession Number – NP_056444. LocusLink ID - 26121.

Properties

Applications

Our Abpromise guarantee covers the use of ab2467 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 Use a concentration of 0.3 - 1 µg/ml. Detects a band of approximately 62 kDa (predicted molecular weight: 55 kDa).Can be blocked with Human PRPF31 peptide (ab22909).

Target

  • Function
    Involved in pre-mRNA splicing. Required for U4/U6.U5 tri-snRNP formation.
  • Tissue specificity
    Ubiquitously expressed.
  • Involvement in disease
    Defects in PRPF31 are the cause of retinitis pigmentosa type 11 (RP11) [MIM:600138]. RP leads to degeneration of retinal photoreceptor cells. Patients typically have night vision blindness and loss of midperipheral visual field. As their condition progresses, they lose their far peripheral visual field and eventually central vision as well. RP11 inheritance is autosomal dominant.
  • Sequence similarities
    Belongs to the PRP31 family.
    Contains 1 Nop domain.
  • Domain
    Interacts with the snRNP via the Nop domain.
    The coiled coil domain is formed by two non-contiguous helices.
  • Cellular localization
    Nucleus speckle. Nucleus > Cajal body. Predominantly found in speckles and in Cajal bodies.
  • Information by UniProt
  • Database links
  • Alternative names
    • DKFZp566J153 antibody
    • hPrp 31 antibody
    • hPrp31 antibody
    • NY BR 99 antibody
    • Pre mRNA processing factor 31 antibody
    • Pre mRNA processing factor 31 homolog (yeast) antibody
    • Pre mRNA processing factor 31 homolog antibody
    • Pre-mRNA-processing factor 31 antibody
    • Precursor mRNA-processing factor 31, S. cerevisiae, homolog of antibody
    • Protein 61K antibody
    • PRP 31 antibody
    • PRP31 antibody
    • PRP31 pre mRNA processing factor 31 homolog (yeast) antibody
    • PRP31 pre mRNA processing factor 31 homolog antibody
    • PRP31_HUMAN antibody
    • PRPF 31 antibody
    • prpf31 antibody
    • RP 11 antibody
    • RP11 antibody
    • Serologically defined breast cancer antigen NY BR 99 antibody
    • Serologically defined breast cancer antigen NY-BR-99 antibody
    • SNRNP61 antibody
    • U4/U6 small nuclear ribonucleoprotein Prp31 antibody
    • U4/U6 snRNP 61 kDa protein antibody
    see all

References

ab2467 has not yet been referenced specifically in any publications.

Customer reviews and Q&As

1-3 of 3 Abreviews or Q&A

Answer

Thank you for your enquiry. I am sorry to hear that you have been having difficulties with this antibody. It is a good selling antibody that we have received very few complaints about. You are performing an approach that I would largely recommend, using the antibody within an appropriate dilution range against a range of protein mass. In your technical questionaire you did not mention how you are preparing your samples and whether you have successfully probed your samples with other antibodies. Please can you detail this, including any protease inhibitors that you have employed. This would certainly be essential in view of the many bands that you have detected. At this stage I would also like to recommend that you try applying the antibody using 3% BSA as a blocking agent instead of milk, applied using an overnight incubation at 4oC. We frequently find that this serves to improve the specificity of the antibody and often provides hugely contrasting results when compared to the use of milk as a blocking agent. I hope this information helps, please do not hesitate to contact us if you need any more advice or information.

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Answer

I'm sorry to hear you are experiencing problems with ab2467. I have asked our laboratory and we have not received any other complaints about this antibody, in fact you received the same master stock in each order. Our policy is to replace or refund if the antibody does not work as described on the datasheet and this guarantee is for 3 months following the purchase of the product. As the product was bought last summer the Abpromise has expired however as a gesture of good will as you are a loyal customer I have exceptionally arranged for a free vial to be sent to you on order 124484. Could you please submit a review with your positive feedback on this antibody so other users can benefit from your experience? We reward each Abreview with 50 Abpoints (and another 100 points for an image), which you can use for future discounts, conference travel money and gifts. I wish you all the best with your research,

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Question

ANTIBODY CODE ab2467 BATCH NUMBER 32490 ORDER NUMBER -- NOT SPECIFIED -- DESCRIPTION OF THE PROBLEM No bands on Western, very high background over entire membrane. SAMPLE Soluble protein extract from transfected 293T cells over-expressing human PRPF31 (His-tagged). Expression of PRPF31-His previously checked with anti-His antibody. PRIMARY ANTIBODY anti human PRPF31 (Abcam2467, lot 23490) used at 2 microgram/ml in PBS 1 hour incubation. Washed 4 x 10min with PBS/Tween SECONDARY ANTIBODY anti-goat IgG HRP conjugate (Abcam6741, lot 34812) used at 1 in 5000 in 10% dried milk powder/PBS 1 hour incubation Washed 2 x 10min with PBS/Tween, then 2 x 10min with PBS only DETECTION METHOD ECLPlus POSITIVE AND NEGATIVE CONTROLS USED Cells transfected with vector only (should express endogenous PRPF31 but at much lower level). ANTIBODY STORAGE CONDITIONS -20deg C, thawed once SAMPLE PREPARATION Hepes buffer containing DTT and PMSF (plus standard SDS-PAGE gel loading buffer). AMOUNT OF PROTEIN LOADED 200 microgram soluble cell protein, amt PRPF31 not estimated ELECTROPHORESIS/GEL CONDITIONS SDS-PAGE, 12% gel TRANSFER AND BLOCKING CONDITIONS Transfer: Zetaprobe membrane Tris/glycine/SDS pH 9.1 30 min transfer time using semi-dry system Blocking: 5% dried milk in PBS/Tween HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 1 HAVE YOU RUN A "NO PRIMARY" CONTROL? No WHAT STEPS HAVE YOU ALTERED? Performed dot blot with same samples, using Hybond C extra and Zetaprobe membranes. Same high background as before using both membranes.Clear patches corresponding to dots of proteins loaded.

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Answer

I am sorry to hear that you are experiencing trouble with this antibody. Zetaprobe is a Biorad nylon membrane recommended for DNA/RNA blotting - not Westerns or any protein applications. Unless there is another product called Zetaprobe this could be behind your background problems. I would strongly recommend using a PVDF membrane. I also recommend doing a no primary control - omit the primary antibody and test with the secondary only; if background appears, change either the secondary or blocking agent. In addition, 200 ug is a high amount to load onto the gel (30-50 ug is generally standard). Here is the protocol which the originator of ab2467 used which I hope will be useful: - Lysis. Cell pellets were washed with ice-cold PBS. 1 ml of RIPA buffer was added per 1E8 cells and incubated on ice for 20 min, vortexing 2-3 times, briefly. The lysate was aliquotted into 1.5 ml microfuge tubes and centrifuged at 13,000 rpm for 5 min in a microfuge. The supernatant was transferred into clean tubes and its protein concentration was measured with BioRad protein assay. The concentration was then adjusted to 5 mg/ml with RIPA lysis buffer. An equal volume of 2 x SDS sample buffer was then added and the cell lysate was boiled for 5 minutes. Lysates were stored at -80C until use.(RIPA buffer = 50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1 mM PMSF, 1 mM EDTA, 5 µg/ml Aprotinin, 5 µg/ml Leupeptin, 1% Triton -100, 1% Sodium deoxycholate, 0.1% SDS). - SDS PAGE. Samples were run at 200V constant on a 12% acrylamide SDS-PAGE mini gel - using Biorad Mini-Protean 3 kit and protocols. Before loading samples had 5% (v/v) 2-ME added and were boiled for 3 minutes. - Transfer. We used a Biorad Mini Trans-Blot, constant 100 V for 1 hour. Transfer Buffer was 20 mM Tris pH 8.0, 150 mM Glycine, 10% Methanol. We transferred to Millipore PVDF membrane and stained with Ponceau Red to evaluate the transfer. - Detection using ECLplus (Amersham) and autoradiography film as described by the manufacturer

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