Nanoparticle-Based Proximity Ligation Assay for Ultrasensitive, Quantitative Detection of Protein Biomarkers

Proximity ligation assay (PLA) achieves extremely low limits of detection but requires the synthesis of antibody–DNA conjugates recognizing multiple target epitopes with appropriate proximity. In this work, we introduce a more generally applicable method by replacing antibody–DNA conjugates with nanoparticles which create ultradetectable PCR templates by capturing biotinylated oligonucleotides and catalyzing ligation. A competitive PCR readout was used to make the assay quantitative. We have demonstrated that NP-PLA can detect and quantitate human chorionic gonadotropin (hCG) levels as low as 2.6 fM (∼0.1 pg/mL), nearly 1000 times more sensitive than the current state of the art ELISA