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NRD1 antibody (Nardilysin (N-Arginine Dibasic Convertase)) (Middle Region) Primary Antibody

NRD1 Reactivity: Cow, Horse, Human, Pig WB Host: Rabbit Polyclonal
Independent Validation (1)
Catalog No. ABIN2787813
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  • Target
    Binding Specificity
    Middle Region
    Reactivity
    Cow, Horse, Human, Pig
    Host
    Rabbit
    Clonality
    Polyclonal
    Conjugate
    This NRD1 antibody is un-conjugated
    Application
    Western Blotting (WB)
    Blocking Peptide
    Blocking peptide for this product available: ABIN980953
    Sequence
    GSKMLSVHVV GYGKYELEED GTPSSEDSNS SCEVMQLTYL PTSPLLADCI
    Predicted Reactivity
    Cow: 93%, Horse: 86%, Human: 100%, Pig: 93%
    Characteristics
    This is a rabbit polyclonal antibody against NRD1. It was validated on Western Blot using a cell lysate as a positive control.
    Purification
    Affinity Purified
    Immunogen
    The immunogen is a synthetic peptide directed towards the middle region of human NRD1
  • Application Notes
    Optimal working dilutions should be determined experimentally by the investigator.
    Comment

    Antigen size: 1151 AA

    Restrictions
    For Research Use only
  • Validation #103724 (Immunohistochemistry)
    'Independent Validation' Badge
    by
    AG Deeg, Lehrstuhl für Physiologie, Veterinärwissenschaftliches Department, Tierärztliche Fakultät, Ludwig-Maximilians-Universität München
    No.
    #103724
    Date
    07/19/2019
    Antigen
    NRD1
    Lot Number
    QC27244-100203
    Method validated
    Immunohistochemistry
    Positive Control

    retina from horses suffering from equine recurrent uveitis (ERU)

    Negative Control

    retina from non-ERU infected horses

    isolated equine retinal Müller glial cells (eqMCs)

    Notes

    Passed. The NRD1 antibody ABIN2787813 specifically labels the targeted antigen in retinae from horses suffering from ERU and in isolated equine retinal Müller glial cells.

    'Independent Validation' Badge
    Validation Images
    Full Methods
    Primary Antibody
    ABIN5074774
    Secondary Antibody
    goat anti-rabbit IgG (H+L) AF568-conjugated antibody (Invitrogen, A11036, lot 1832035)
    Full Protocol
    • IHC on fixed equine retina:
      • Dissect tissue and fix in 4% buffered formalin for 12h at 4°C.
      • Dehydrate and paraffinize the tissue through a graded alcohole, xylene and paraffine series.
      • Embed the tissue in Richard Allen Scientific Paraffine Type 9 (Thermofisher, 8337).
      • Cut fixed tissue with a microtome into 8µm-thick sections.
      • Deparaffinization and rehydration through graded xylene and graded alcohol series:
      • Boil the sections in a 99°C hot waterbath in 0.1M Na2-EDTA buffer pH8.0 for 15min.
      • Boil the sections in a 99°C hot waterbath in citrate buffer pH6 (Dako, 2031) for 15min.
      • Rinse sections with PBS pH7.4.
    • IHC on isolated eqMCs:
      • Prepare eqMCs from the eyes of a 19 year old mare.
      • Cultivate cells in DMEM (PAN-Biotech, P04-04510, lot 2650219) supplemented with 10% v/V FBS (Biochrom, S0615, lot 0879F) and 1% Penicillin/Streptomycin (PAN-Biotech, P06-07100, lot 7180417) for 18d at 37°C at 5% CO2.
      • Seed 1x105 cells (first passage) in 1ml medium on adhesion microscope slides (Langenbrinck 03-0060, lot 021218).
      • Incubate cells for 2d at 37°C at 5% CO2.
      • Remove medium and wash cells with PBS pH7.4.
      • Fix cells for 10min with ice-cold acetone.
      • Rehydrate cells with TBST (0.01M Tris, 0.15M NaCl, 0.1% v/v Tween 20) pH7.3.
    • Block samples in 5% goat serum in TBST pH7.3 containing 1% w/v BSA (BSA-TBST).
    • Incubate samples with primary rabbit anti-NRD1 antibody (antibodies-online, ABIN2787813, lot QC27244-100203) diluted 1:100 in BSA-TBST ON at 4°C.
    • Wash samples with BSA-TBST.
    • Incubate samples with secondary goat anti-rabbit IgG (H+L) AF568-conjugated antibody (Invitrogen, A11036, lot 1832035) diluted 1:500 in BSA-TBST for 30min at RT.
    • Wash samples with BSA-TBST.
    • Incubate samples with primary mouse anti-Vimentin antibody (Sigma-Aldrich, V6630) diluted 1:400 in BSA-TBST ON at 4°C.
    • Wash samples with BSA-TBST.
    • Incubate samples with secondary goat anti-mouse IgG (H+L) AF488-conjugated antibody (Invitrogen, A11029, lot 1874804) diluted 1:500 in BSA-TBST containing 0.1% DAPI for 30min at RT.
    • Wash samples with BSA-TBST.
    • Mount samples in Fluoromount W mounting medium (SERVA, 21634, lot 181008).
    • Acquire images with Leica DMi8 fluorescence microscope equipped with Filter Cubes for DAPI, FITC, TXR and Y5 at 200x (prepared retina) and 400x (eqMCs). Parameters for image acquisition (exposure: FITC 220ms, TXR 253ms, DAPI 195ms) were maintained unchanged for all images.
    Experimental Notes

    ABIN2787813 was also tested in western blot on retinal Müller glial cell lysate from healthy horses or horses suffering from ERU and in equine and porcine total retinal lysates. However, the antibody did not detect a protein of the expected MW of 132kDa in any of these samples whereas numerous extraneous protein bands were detected.

  • Format
    Liquid
    Concentration
    Lot specific
    Buffer
    Liquid. Purified antibody supplied in 1x PBS buffer with 0.09 % (w/v) sodium azide and 2 % sucrose.
    Preservative
    Sodium azide
    Precaution of Use
    This product contains Sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.
    Handling Advice
    Avoid repeated freeze-thaw cycles.
    Storage
    -20 °C
    Storage Comment
    For short term use, store at 2-8°C up to 1 week. For long term storage, store at -20°C in small aliquots to prevent freeze-thaw cycles.
  • Target
    Alternative Name
    NRD1 (NRD1 Antibody Abstract)
    Background
    NRD1 cleaves peptide substrates on the N-terminus of arginine residues in dibasic pairs.
    Alias Symbols: hNRD1, hNRD2
    Protein Interaction Partner: FBXW11, UBC, rev, SETSIP, PUS1, P3H1, ELAC2, XPO5, NSUN2, PDIA6, YARS, DNAJC7, TSN, PPP3CA, PPP2R2A, PPP2CA, PFDN5, PFDN1, PPP1R12A, MSH2, PNKP, HSPB1, MOB2, MOB1A, STK3, NRD1, HIST2H3C, UACA, COBLL1, NCOR2, NCOR1, HDAC3, IQGAP1, MAD1L1, TBL1X, UIMC1, BTR
    Protein Size: 1151
    Molecular Weight
    132 kDa
    Gene ID
    4898
    NCBI Accession
    NM_001101662, NP_001095132
    UniProt
    O43847
    Pathways
    Skeletal Muscle Fiber Development
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