A platform for ultrasensitive and selective multiplexed marker protein assay toward early-stage cancer diagnosis

Multiplexed marker protein assay is critical in the diagnosis of complex diseases that cannot be diagnosed by detection of a single marker protein. Gold nanoparticle (Au NP) probes barcoded with reporter DNAs and magnetic microparticles functionalized with a capture antibody were developed for the multiplexed detection of three cancer marker proteins. Three types of Au NP probes were used and each Au NP probe was cofunctionalized with reporter (barcode) DNAs and a specific antibody for each corresponding target protein. Target proteins (antigens) were exposed to magnetic microparticles to form complexes and Au NP probes bound to the resulting target-magnetic microparticle complex through antigen-antibody interaction in a different region of the target protein. After magnetic separation of the complexes, barcode DNAs were released, hybridized with capture DNAs printed on a chip and then identified using a scanometric assay that involved silver amplification. Using this method, Mirkin and colleagues successfully demonstrated, for the first time, a highly selective and sensitive multiplexed protein assay against three cancer marker proteins at low picomolar concentration in a buffer of serum media.