A rapid and sensitive method to detect cardiac troponin I (cTnI) in human blood is critical to the diagnosis and treatment of acute myocardial infarction (AMI). Here, we describe a simple one-step digital immunoassay for single-molecule detection without washing steps. A sample containing cTnI mixed with detection antibody-conjugated gold nanoparticles (AuNPs) is added to a capture antibody-coated sensor surface and the formation of the antibody-cTnI-antibody sandwich is detected by digitally counting the binding of the individual gold nanoparticles to the sensor surface in real time using a bright-field optical imaging setup together with a differential imaging algorithm. The digital immunoassay detects cTnI in undiluted human plasma, which achieves a detection limit of 5.7 ng/L within a detection time of only 10 min, which meets the requirement of current clinical high-sensitivity troponin assay (∼70 ng/L cutoff). We anticipate that the one-step and real-time digital immunoassay can be applied to the detection of other disease biomarkers in blood.
Keywords: cardiac troponin I; digital immunoassay; high-sensitivity troponin assay; one-step immunoassay; single nanoparticle counting.