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Abstract

Aim: Nowadays, IFN-α is considered a promising therapeutic target for systemic lupus erythematosus. An immuno-PCR (iPCR) was developed to quantify low amounts of IFN-α in human plasma followed by a deep analysis of the methodologic robustness throughout quality by design approach. Results: An accurate, sensitive, selective and versatile iPCR was validated. The critical iPCR procedural steps were identified, applying a Plackett–Burman design. Also, this assay demonstrated an outstanding LOD of 0.3 pg/ml. A significant aspect relies on its high versatility to detect and quantify other cytokines in human plasma as the appropriate biotinylated antibody is employed. Conclusion: This reliable iPCR assay can be clinically used as an alternative method for quantitating and detecting low IFN-α2b concentrations in human plasma samples.

Graphical abstract

Keywords:

  • accuracy
  • ELISA
  • IFN-α
  • immuno-PCR
  • linear range
  • precision
  • quality by design
  • robustness
  • selectivity
  • sensitivity
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