Human PU.1/Spi-1 Antibody

  
  • Species Reactivity
    Human
  • Specificity
    Detects human PU.1 in direct ELISAs and Western blots. In direct ELISAs, approximately 5% cross-reactivity with recombinant human SPI-B is observed.
  • Source
    Polyclonal Sheep IgG
  • Purification
    Antigen Affinity-purified
  • Immunogen
    E. coli-derived recombinant human PU.1
    Met1-Lys169
    Accession # NP_001074016
  • Formulation
    Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied as a 0.2 µm filtered solution in PBS.
  • Label
    Unconjugated
Applications
  •  
    Recommended
    Concentration
    Sample
  • Western Blot
    1 µg/mL
    See below
  • CyTOF-ready
    Ready to be labeled using established conjugation methods. No BSA or other carrier proteins that could interfere with conjugation.
  • Immunocytochemistry
    5-15 µg/mL
    See below
  • Intracellular Staining by Flow Cytometry
    2.5 µg/106 cells
    PMA and Ca2+ ionomycin‑treated human Th2-stimulated PBMCs, fixed with paraformaldehyde, and permeabilized with saponin
Please Note: Optimal dilutions should be determined by each laboratory for each application. General Protocols are available in the Technical Information section on our website.
Data Examples
Detection of Human PU.1 by Western Blot. Western blot shows lysates of THP‑1 human acute monocytic leukemia cell line. PVDF membrane was probed with 1 µg/mL of Human PU.1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF5870) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). A specific band was detected for PU.1 at approximately 45 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 8.
Immunocytochemistry
PU.1 in THP‑1 Human Cell Line. PU.1 was detected in immersion fixed THP‑1 human acute monocytic leukemia cell line using Human PU.1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF5870) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Sheep IgG Secondary Antibody (red; Catalog # NL010) and counterstained with DAPI (blue). Specific staining was localized to nuclei and cytoplasm. View our protocol for Fluorescent ICC Staining of Non-adherent Cells.
Preparation and Storage
  • Reconstitution
    Reconstitute at 0.2 mg/mL in sterile PBS.
  • Shipping
    The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below. *Small pack size (SP) is shipped with polar packs. Upon receipt, store it immediately at -20 to -70 °C
  • Stability & Storage
    Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
    • 12 months from date of receipt, -20 to -70 °C as supplied.
    • 1 month, 2 to 8 °C under sterile conditions after reconstitution.
    • 6 months, -20 to -70 °C under sterile conditions after reconstitution.
Background: PU.1/Spi-1
PU.1 (Purine-rich nucleic acid binding protein 1; also 31 kDa transforming protein and SPI-1) is a member of the PU subfamily, ETS family of transcription factors. Although its predicted MW is 31 kDa, it appears to run anomalously high in SDS-PAGE at 40‑45 kDa. PU.1 is a monomeric hematopoietic protein that regulates the differentiation of early myeloid and lymphoid progenitors. High PU.1 levels favor granulocyte and macrophage production, while low levels generate magakaryocytes, erythrocytes, T and B cells. Human PU.1 is 270 amino acids (aa) in length. It contains an N-terminal acidic/polyGln transactivation region (aa 34‑99), a non‑destabilizing PEST sequence (aa 117‑165) and a C-terminal Ets DNA-binding domain (aa 170‑253). PU.1 is phosphorylated on Ser146, allowing it to bind to Pip. Over aa 1‑169, human PU.1 shares 88% aa identity with mouse PU.1.
  • Long Name:
    Hematopoietic transcription factor PU.1
  • Entrez Gene IDs:
    6688 (Human); 20375 (Mouse); 366126 (Rat)
  • Alternate Names:
    hematopoietic transcription factor PU.1; OF; PU.1; SFPI1; SFPI131 kDa transforming protein; SPI-1 proto-oncogene; SPI1; Spi-1; SPI-A; spleen focus forming virus (SFFV) proviral integration oncogene spi1,31 kDa-transforming protein; transcription factor PU.1
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