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Testing NF-kappa B1-p50 antibody specificity using knockout mice

D S Pereira1, D B Kushner, R P Ricciardi

  • 1Department of Biology, McMaster University, Hamilton, Ontario, Canada.

Oncogene
|July 18, 1996
PubMed
Summary
This summary is machine-generated.

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Antibody specificity for NF-kappa B1-p50 was tested using knockout mice. NR1157 antibody showed high specificity, while sc-114 did not, highlighting the utility of knockout models.

Area of Science:

  • Immunology
  • Molecular Biology
  • Biochemistry

Background:

  • Antibody specificity is crucial for accurate Western blot analysis.
  • NF-kappa B1-p50 is a key transcription factor involved in immune responses.
  • Distinguishing between specific and non-specific antibody binding is essential for reliable experimental results.

Purpose of the Study:

  • To determine the specificity of two antibodies, sc-114 and NR1157, against NF-kappa B1-p50.
  • To evaluate the utility of NF-kappa B1-p105 knockout mouse cell extracts for antibody validation.

Main Methods:

  • Western blot analysis was performed using two antibodies (sc-114 and NR1157) against whole cell extracts.
  • NF-kappa B1-p105 knockout mouse cell extracts (p105-/-) were utilized to assess antibody specificity.

Related Experiment Videos

  • Electrophoretic mobilities of recognized proteins were compared between wild-type and knockout conditions.

    • Main Results:

    • Antibody sc-114 recognized a 52-55 kD protein in both wild-type and knockout extracts.
    • Antibody NR1157 recognized a 50 kD protein in wild-type extracts but failed to detect it in knockout extracts.
    • These findings indicate that NR1157 is specific for NF-kappa B1-p50, whereas sc-114 shows cross-reactivity.

    Conclusions:

    • The NR1157 antibody demonstrates high specificity for NF-kappa B1-p50 via Western blot.
    • NF-kappa B1-p105 knockout cell extracts are a valuable tool for confirming antibody specificity.
    • This study validates NR1157 as a specific reagent for detecting NF-kappa B1-p50.