Selective detection of biomarkers at low concentrations in blood is crucial for the clinical diagnosis of many diseases but remains challenging. In this work, we aimed to develop an ultrasensitive immunoassay that can detect biomarkers in serum with an attomolar limit of detection (LOD). We proposed a sandwich-type heterogeneous immunosensor in a 3 × 3 well array format by integrating a resonant waveguide grating (RWG) substrate with upconversion nanoparticles (UCNPs). UCNPs were used to label a target biomarker captured by capture antibody molecules immobilized on the surface of the RWG substrate, and the RWG substrate was used to enhance the upconversion luminescence (UCL) of UCNPs through excitation resonance. The LOD of the immunosensor was greatly reduced due to the increased UCL of UCNPs and the reduction of nonspecific adsorption of detection antibody-conjugated UCNPs on the RWG substrate surface by coating the RWG substrate surface with a carboxymethyl dextran layer. The immunosensor exhibited an extremely low LOD [0.24 fg/mL (9.1 aM)] and wide detection range (1 fg/mL to 100 pg/mL) in the detection of cardiac troponin I (cTnI). The cTnI concentrations in human serum samples collected at different times during cyclophosphamide, epirubicin, and 5-fluorouracil (CEF) chemotherapy in a breast cancer patient were measured by an immunosensor, and the results showed that the CEF chemotherapy did cause cardiotoxicity in the patient. Having a higher number of wells in such an array-based biosensor, the sensor can be developed as a high-throughput diagnostic tool for clinically important biomarkers.
Keywords: cardiac troponin I; excitation resonance; guided mode resonance; resonant waveguide grating; sandwich-type heterogeneous immunoassay; upconversion luminescence; upconversion nanoparticle.