IgG-detection devices for the Tus-Ter-lock immuno-PCR diagnostic platform

Area of Science:

• Biotechnology
• Assay Development
• Molecular Diagnostics

Background:

• Immuno-PCR (IPCR) offers superior sensitivity over traditional immunoassays but faces challenges like high background noise and poor reproducibility.
• Developing homogeneous, universal protein-DNA conjugates is crucial for simplifying IPCR and improving its robustness.

Purpose of the Study:

• To further develop the Tus-Ter-lock (TT-lock) Immuno-PCR platform for quasi-universal quantitative detection of antigens and mammalian IgG.
• To enhance the sensitivity and reproducibility of Immuno-PCR assays for early disease detection.

Main Methods:

• Engineered Tus protein fusions with IgG-binding proteins (Protein G, Protein L, LG chimera) to create homogeneous protein-DNA conjugates.
• Utilized the TT-lock interaction for self-assembly of detection devices on a TT-lock-T template.
• Applied direct and indirect TT-lock Immuno-PCR formats for quantitative detection of antibodies and antigens.

Main Results:

• Direct TT-lock qIPCR detected goat anti-GFP IgG at 0.3 pM and human IgG in serum with high sensitivity.
• Indirect TT-lock qIPCR systems detected 1 pM GFP and 10 pM measles nucleoprotein.
• Demonstrated superior sensitivity of TT-lock Immuno-PCR compared to Protein G-Peroxidase ELISA.

Conclusions:

• The developed TT-lock Immuno-PCR platform provides a robust and highly sensitive method for quantitative detection of various disease biomarkers.
• This technology simplifies Immuno-PCR, addressing key limitations and paving the way for improved early disease diagnostics.