Recombinant
RabMAb

Anti-PUS1 antibody [EPR20181] (ab203010)

Overview

  • Product name
    Anti-PUS1 antibody [EPR20181]
    See all PUS1 primary antibodies
  • Description
    Rabbit monoclonal [EPR20181] to PUS1
  • Tested applications
    Suitable for: Flow Cyt, WB, ICC/IF, IPmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    Recombinant fragment within Human PUS1 aa 50 to the C-terminus. The exact sequence is proprietary.
    Database link: Q9Y606

  • Positive control
    • WB: Human skeletal muscle lysate; HeLa mitchodrial and cytoplasm fraction lysates; A431, HEK-293, C6, RAW 264.7, PC-12 and NIH/3T3 whole cell lysates; Mouse heart and spleen lysates; Rat spleen lysate. ICC/IF: HeLa cells. Flow Cyt: HeLa cells. IP: HEK-293 whole cell lysate.
  • General notes

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab203010 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
Flow Cyt 1/400.
WB 1/1000. Detects a band of approximately 47, 44 kDa (predicted molecular weight: 47 kDa).
ICC/IF 1/500.

This antibody was not successful when we used it on RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) cells in ICC application. This antibody was not tested on rat cells in ICC.

IP 1/30.

Target

  • Function
    Converts specific uridines to PSI in a number of tRNA substrates. Acts on positions 27/28 in the anticodon stem and also positions 34 and 36 in the anticodon of an intron containing tRNA. Involved in regulation of nuclear receptor activity possibly through pseudouridylation of SRA1 RNA.
  • Tissue specificity
    Widely expressed. High levels of expression found in brain and skeletal muscle.
  • Involvement in disease
    Defects in PUS1 are a cause of myopathy with lactic acidosis and sideroblastic anemia type 1 (MLASA1) [MIM:600462]; also known as mitochondrial myopathy and sideroblastic anemia. MLASA is a rare autosomal recessive oxidative phosphorylation disorder specific to skeletal muscle and bone marrow.
  • Sequence similarities
    Belongs to the tRNA pseudouridine synthase TruA family.
  • Cellular localization
    Mitochondrion and Nucleus.
  • Information by UniProt
  • Database links
  • Alternative names
    • A730013B20Rik antibody
    • DOBI antibody
    • MGC112655 antibody
    • MGC11268 antibody
    • MLASA antibody
    • MLASA1 antibody
    • mPus1p antibody
    • Pseudouridine synthase 1 antibody
    • Pseudouridylate synthase 1 antibody
    • PUS1 antibody
    • tRNA pseudouridine synthase A, mitochondrial antibody
    • tRNA pseudouridine(38-40) synthase antibody
    • tRNA pseudouridylate synthase I antibody
    • tRNA uridine isomerase I antibody
    • tRNA-uridine isomerase I antibody
    • TRUA_HUMAN antibody
    see all

Images

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling PUS1 with ab203010 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining on HeLa cell line.

    The nuclear counterstain is DAPI (blue). Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594)) at 1/200 dilution (red).

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) at 1/1000 dilution.

    This antibody was not successful when we used it on RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) cells in ICC application. This antibody was not tested on rat cells in ICC.

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling PUS1 with ab203010 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing mitochondrial staining on HeLa cell line.

    The nuclear counterstain is DAPI (blue). COXIV is detected with ab33985 (Anti-COX IV(mouse mAb)) at 1/1000 dilution followed by Goat anti-mouse IgG (Alexa Fluor® 594) (ab150120) secondary antibody at 1/1000 dilution (red).

    The negative controls are as follows:-

    -ve control 1: ab203010 at 1/500 dilution followed by ab150120 (Alexa Fluor® 594 Goat anti-Mouse secondary) at 1/1000 dilution.

    -ve control 2: ab33985 (anti-COX IV(mouse mAb)) at 1/1000 dilution followed by ab150077 (Alexa Fluor® 488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.

  • Anti-PUS1 antibody [EPR20181] (ab203010) at 1/1000 dilution + Human skeletal muscle lysate at 10 µg

    Secondary
    Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/1000 dilution

    Predicted band size : 47 kDa
    Observed band size : 47 kDa


    Exposure time : 3 minutes

    Blocking/Dilution buffer: 5% NFDM/TBST.

  • All lanes : Anti-PUS1 antibody [EPR20181] (ab203010) at 1/5000 dilution

    Lane 1 : HeLa (Human epithelial cell line from cervix adenocarcinoma) mitochondria lysate
    Lane 2 : HeLa cytoplasm fraction lysate
    Lane 3 : A431 (Human epidermoid carcinoma cell line) whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size : 47 kDa
    Observed band size : 47 kDa


    Exposure time : 8 seconds

    Blocking/Dilution buffer: 5% NFDM/TBST.

    The expression profile observed is consistent with what has been described in UniProt.

  • Anti-PUS1 antibody [EPR20181] (ab203010) at 1/1000 dilution + HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate at 10 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size : 47 kDa
    Observed band size : 44,47 kDa (why is the actual band size different from the predicted?)


    Exposure time : 3 seconds

    Blocking/Dilution buffer: 5% NFDM/TBST.

    Based on sequence alignment, the antibody can recognize 2 isoforms, the predicted MW are 47kDa and 44kDa, respectively [PMID: 17056637].

  • All lanes : Anti-PUS1 antibody [EPR20181] (ab203010) at 1/2000 dilution

    Lane 1 : Mouse heart lysate
    Lane 2 : Mouse spleen lysate
    Lane 3 : Rat spleen lysate
    Lane 4 : C6 (Rat glial tumor cell line) whole cell lysate
    Lane 5 : RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate
    Lane 6 : PC-12 (Rat adrenal gland pheochromocytoma cell line) whole cell lysate
    Lane 7 : NIH/3T3 (Mouse embryonic fibroblast cell line) whole cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size : 47 kDa
    Observed band size : 47 kDa

    Blocking/Dilution buffer: 5% NFDM/TBST.

    Exposure time: Lane 1 and 2: 10 seconds; Lane 3: 3minutes; Lane 4,5,6 and 7: 10 seconds.

  • Flow cytometric analysis of 4% paraformaldehyde-fixed HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling PUS1 with ab203010 at 1/400 dilution (red) compared with a rabbit monoclonal IgG isotype control (ab172730; black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (Alexa Fluor® 488) at 1/2000 dilution was used as the secondary antibody.

  • PUS1 was immunoprecipitated from 0.35 mg of HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate with ab203010 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab203010 at 1/1000 dilution. VeriBlot for IP secondary antibody (HRP) (ab131366), was used as secondary antibody at 1/10000 dilution.

    Lane 1: HEK-293 whole cell lysate, 10µg (Input).

    Lane 2: ab203010 IP in HEK-293 whole cell lysate.

    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab203010 in HEK-293 whole cell lysate.

    Blocking and dilution buffer and concentration: 5% NFDM/TBST.

    Exposure time: 5 seconds.

References

ab203010 has not yet been referenced specifically in any publications.

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