Highly sensitive detection of cardiac troponin i in human serum using gold nanoparticle-based enhanced sandwich immunoassay

We have developed an enhanced sandwich immunogold assay for highly sensitive detection of cardiac troponin I (cTnI) in human serum. In this assay, the immunoreaction undergoes in sample solution between cTnI antigen and gold nanoparticles conjugated with detection antibodies of cTnI (immunogolds). Then, a capture antibody of cTnI immobilized on a standard 96-well plate captures the immunogold complex, followed by silver staining of the immunogold for the purpose of signal amplification. Compared to the direct assay where the immunoreaction occurs between the target and the capture antibody coated on the well, we found the enhanced assay shows greatly improved sensitivity by two orders of magnitude and a linear relation of the quantitative plot. Because the immunogold concentration affect greatly the sensitivity of the cTnI detection, we also studied the performance of direct and enhanced immunogold assay depending on the concentration of immunogold conjugates. By using the enhanced assay with the optimized procedure, we could achieve the detection of cTnI in human serum as low as 15 pg/mL. We also verified the enhanced immunogold assay by measuring the concentration of clinical samples for a healthy person and patients. This development can allow us to apply the assay not only for the analysis of other cardiac markers as well as disease-related biomarkers but also for the optimization of other chip-based silver enhanced colorimetric sensors.