Immunoassays with protein misfolding cycle amplification: a platform for ultrasensitive detection of antigen

Protein misfolding cycle amplification (PMCA), a novel technology on amplifying cyclically misfolded proteins in vitro, is conceptually analogous to DNA amplification by polymerase chain reaction (PCR) and has tremendous implications for the researches and diagnosis. Here we first introduce the protein amplification technology into the classic immunoassay and develop a PMCA-based immunoassay (immuno-PMCA) for highly sensitive detection of antigen. This method takes advantage of sandwich binding of two affinity aptamers for increased specificity, magnetic nanoparticles for fast magnetic separation, PMCA for signal amplification, and conjugated polyelectrolytes for visual detection, allowing the detection limit of antigen by colorimetry down to femtomolar level with a wide linear range from 10 to 10(4) fM. More importantly, no specialized facilities or enzymes are needed either in the amplification reaction or the evaluation of results, which indicates its great potential application in immunological research and clinical diagnostics.