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GST antibody (Glutathione S Transferase) Primary Antibody

GST Reactivity: Schistosoma japonicum IP, WB Host: Mouse Monoclonal 6C10G4
Independent Validation (1)
Catalog No. ABIN1998462
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  • Target
    Reactivity
    Schistosoma japonicum
    Host
    Mouse
    Clonality
    Monoclonal
    Conjugate
    This GST antibody is un-conjugated
    Application
    Immunoprecipitation (IP), Western Blotting (WB)
    Specificity
    Recognize GST Tag in fusion proteins.
    No cross-reactivity with E.coli cell lysate in ELISA.
    No Cross-Reactivity
    Escherichia coli (E. coli)
    Characteristics
    This antibody was produced from a hybridoma resulting from the fusion of a mouse myeloma with B cells obtained from a mouse immunized with purified, recombinant Glutathione S-transferase (GST). The IgG fraction of the cell culture supernatant was purified by Protein A affinity chromatography.
    Purification
    purified by Protein A affinity chromatography
    Sterility
    0.2 μm filtered
    Immunogen
    Recombinant GST protein
    Clone
    6C10G4
    Isotype
    IgG1
  • Target
    Alternative Name
    GST (GST Antibody Abstract)
    Background
    Genetic engineers have used glutathione S-transferase to create the GST gene fusion system. This system is used to purify and detect proteins of interest. In a GST gene fusion system, the GST sequence is incorporated into an expression vector alongside the gene sequence encoding the protein of interest. Induction of protein expression from the vector's promoter results in expression of a fusion protein: the protein of interest fused to the GST protein. This GST-fusion protein can then be purified from cells via its high affinity for glutathione. GST is commonly used to create fusion proteins. The tag has the size of 220 amino acids (roughly 26 KDa), which, compared to other tags like the myc- or the FLAG-tag, is quite big. However, many commercially-available sources of GST-tagged plasmids include a thrombin domain for cleavage of the GST tag during protein purification.
  • Application Notes
    WB: 1/1000-1/10000
    IP: 1-4 μL/mg of lysate
    Restrictions
    For Research Use only
  • Validation #029849 (Western Blotting)
    'Independent Validation' Badge
    by
    Alamo Laboratories Inc
    No.
    #029849
    Date
    11/06/2015
    Antigen
    Glutathione S Transferase (GST)
    Lot Number
    HD04FE1006
    Method validated
    Western Blotting
    Positive Control
    GST- AcNFkBp(C-terminal) 293-Lysate
    Negative Control
    Empty vector- 293 Lysate
    Notes
    A strong specific band was observed in the positive control at the expected size (~82 kDa) that is not observed in the negative control.
    'Independent Validation' Badge
    Validation Images
    Full Methods
    Primary Antibody
    • Antigen: Glutathione S Transferase (GST) protein
    • Catalog number: ABIN1998462
    • Lot number: HD04FE1006
    • Antibody Dilution: 1:7000
    Secondary Antibody
    • Antigen: Goat Anti-Mouse IgG (H + L)-HRP Conjugate
    • Lot number: N/A
    • Antibody Dilution: 1:15,000
    Full Protocol
    • 1. The cell extracts were heated at 95°C for 5 minutes in 1X SDS Sample Buffer containing 1% SDS and 1.25% β-mercaptoethanol.
    • 2. 15 μl of heated were loaded and resolved on 8-16% SDS-polyacrylamide gel.
    • 3. The Bio-Rad Precision Plus (Cat 161-0374) were used as molecular mass markers.
    • 4. Proteins were then transferred onto PVDF membrane by wet transfer.
    • 5. The PVDF membrane was incubated with 25 ml of blocking buffer [Tris Buffered Saline, pH 7.4 plus 0.1% TW20 (TBST)] containing 5% (W/V) BSA at room temperature for 1 hour.
    • 6. The membrane was rinsed with TBST once.
    • 7. The membrane was immersed with the protein side up in the primary antibody solution in TBST containing 5% (W/V) BSA and incubated for 24 hours at 4°C.
    • 8. The membrane was rinsed in TBST thrice for 5 minutes each.
    • 9. The membrane was incubated in the HRP-conjugated secondary antibody solution in TBST containing 5% (W/V) BSA and incubated for 1 hour at room temperature (~26°C) with gentle agitation.
    • 10. The membrane was rinsed thrice TBST thrice for 5 minutes each.
    • 11. The membrane was rinsed in TBS twice for 30 seconds each.
    • 12. Signals were detected with ECL-2 Substrate. The blot was scanned for 1 minute.
    • 13. The membrane was rinsed three times TBST.
    • 14. Incubated in Acidic Glycine Stripping Buffer at room temperature with gentle agitation for 3 times, 10 minutes each.
    • 15. The membrane was washed in TBST 2 times for 10 minutes each.
    • 16. Repeated Steps 5-12 with the loading control antibody (for Anti-actin) and its matching secondary antibody.
    Experimental Notes
    None reported
  • Format
    Liquid
    Buffer
    0.2 μm filtered solution in PBS
    Preservative
    Without preservative
    Handling Advice
    Avoid repeated freeze-thaw cycles.
    Storage
    4 °C,-20 °C,-80 °C
    Storage Comment
    This antibody can be stored at 2°C-8°C for one month without detectable loss of activity. Antibody products are stable for twelve months from date of receipt when stored at -20°C to -80°C. Preservative-Free.
    Sodium azide is recommended to avoid contamination (final concentration 0.05%-0.1%). It is toxic to cells and should be disposed of properly. Avoid repeated freeze-thaw cycles.
    Expiry Date
    12 months
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