A magnetic electrochemical immunosensor for the detection of phosphorylated p53 based on enzyme functionalized carbon nanospheres with signal amplification

Abstract

Protein phosphorylation plays an important role in many biological processes and might be used as a potential biomarker in clinical diagnoses. We reported the development of a nanomaterial enhanced disposable immunosensor for ultrasensitive detection of phosphorylated p53 at Ser392 (phospho-p53³⁹²) using enzyme functionalization of carbon nanospheres (CNSs) as a signal amplification label and magnetic beads (MB) coupled with screen printed carbon electrodes as electrochemical transducers. In this work, horseradish peroxidase (HRP) and phospho-p53³⁹² detection antibody (Ab₂) were co-linked to CNSs (HRP–CNSs–Ab₂) for signal amplification, and functionalized MB was used as a platform to capture a large amount of primary antibodies (Ab₁). The proposed signal amplification strategy with a sandwich-type immunoreaction significantly enhanced the sensitivity of detection of biomarkers. Under optimal conditions, the immunosensor had a highly linear voltammetric response to the phospho-p53³⁹² concentration in the range of 0.01 to 5 ng mL⁻¹, with a detection limit of 3.3 pg mL⁻¹. The results provided great potential for point-of-care detection of other phosphorylated proteins and clinical applications.