Signaling redox probe/DNA aptamer complexes on a new POP/2D WSe2 composite-based immunosensor towards the simultaneous detection of three-protein overexpression as an alternative severe SARS-COV-2 infection diagnosis

The coronavirus disease 2019 (COVID-19) has become a global pandemic in which patients with symptoms are infected with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Clinical investigation of human immune response cytokines or/and overexpressed proteins at trace cut-off levels can be valuable to monitor body function failures and, eventually, improve the prognosis for patients. Nevertheless, the currently available technologies are still limited. Herein, we provide the first sandwich-like immunosensor that has the potential to detect multiple protein biomarkers in a single measurement. The sandwich-like structure was created by coating a three-screen printed carbon electrode (3SPCE) array with a porous organic polymer (POP)/2D WSe2 nanocomposite. This system enables the simultaneous immunoassay of C-reactive protein (CRP), native cardiac troponin I (cTnI), and procalcitonin (PCT), which together provide a panel of symptom-diagnostic biomarkers for the presence and severity of COVID-19 infection. The biomarker detection principle relies on three antibodies as bioreceptor probes bound on the modified 3SPCE and three aptamers to generate three redox probe complexes as signal amplifiers. After incorporating three redox species (Cd2+, methylene blue (MB), and Ag+) into the aptamer structure, well-differentiated oxidation peaks can be achieved, indicating the concentrations of these three proteins. Under the optimized conditions of the stepwise fabrications, the linear dynamic range of three-protein detection by our proposed multiplex biosensor ranges from 100 to 50,000 pg mL-1. The limits of detection (LODs) are as low as 62, 81, and 44 pg mL-1 for the detection of CRP, cTnI, and PCT, respectively. As proof of the concept, this multiplexed assay demonstrates outstanding analytical performance with excellent reproducibility, high sensitivity and selectivity, and acceptable recoveries of the detection in human serum. Therefore, the multiplexed sensor could be an alternative tool in the field of clinical analysis for the diagnosis of COVID-19 severity or other emerging infectious diseases.