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Related Experiment Videos

Phage-displayed La/SS-B antigen as a diagnostic reagent

V K Poon1, Y L Chui, P L Lim

  • 1Clinical Immunology Unit, Chinese University of Hong Kong, Prince of Wales Hospital.

Immunotechnology : an International Journal of Immunological Engineering
|June 1, 1997
PubMed
Summary

This study developed an easy and inexpensive method to produce La/SS-B antigen using phage display for diagnosing Sjogren's syndrome and SLE. The phage-displayed antigen effectively detected anti-La/SS-B antibodies in patient sera.

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Area of Science:

  • Immunology
  • Molecular Biology
  • Biochemistry

Background:

  • Antibodies to La/SS-B, a nuclear RNA-binding protein, are crucial for diagnosing Sjogren's syndrome and systemic lupus erythematosus (SLE).
  • Conventional detection methods include immunoprecipitation and ELISA using purified La/SS-B antigen, often obtained via affinity chromatography.
  • There is a need for simpler, more cost-effective antigen production methods for diagnostic assays.

Purpose of the Study:

  • To develop an easily and inexpensively produced La/SS-B antigen for ELISA.
  • To avoid extensive purification steps like affinity chromatography for antigen preparation.
  • To utilize phage display technology for antigen production.

Main Methods:

  • La/SS-B cDNA was cloned into a phagemid (pCANTAB-5E) and expressed as a fusion protein with M13 bacteriophage minor coat protein in Escherichia coli.

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  • Phage clones displaying the La/SS-B antigen were selected and used directly in ELISA.
  • The phage-displayed antigen was precipitated from bacterial culture supernatant using polyethylene glycol.

    • Main Results:

    • A phage clone expressing a truncated La/SS-B cDNA fragment (up to base-pair 631) was successfully generated.
    • The phage-displayed La/SS-B antigen detected 27 out of 28 precipitin-positive sera from patients with Sjogren's syndrome and SLE.
    • The assay showed high specificity, with negative results for 50 control sera.
    • A soluble, phage-free form of the antigen was obtained but was only suitable for Western blot analysis, not ELISA.

    Conclusions:

    • Phage-displayed antigens offer a viable and advantageous alternative to soluble antigens in diagnostic detection assays.
    • This method provides an easy and inexpensive approach for producing antigens for immunological assays.
    • Phage display technology simplifies antigen production for diagnosing autoimmune diseases like Sjogren's syndrome and SLE.