Development of Highly Sensitive Label-Free Electrochemical Immunosensor for p16INK4a Detection: A Step Toward Early Cervical Cancer Diagnosis

Abstract

Cervical cancer screening is the key public health approach for secondary prevention of cervical cancer particularly in resource limited countries. This study aims to present development of a highly sensitive electrochemical immunosensor for the detection of p16INK4a, a protein overexpressed in cervical cancer. The immunosensor consisted of a layer-by-layer hybrid film of gold nanoparticles (AuNPs) and reduced graphene oxide (rGO) co-deposited on glassy carbon electrodes (GCE), which can improve electrical conductivity and the effective surface area for immobilizing capture antibody. The anti- p16INK4a monoclonal antibodies were immobilized on the modified electrode surface through a cystamine-glutaraldehyde crosslinking approach. Square wave voltammetry (SWV) technique was employed for p16INK4a detection, exhibiting a limit of detection (LOD) of 167 fg mL-1 and a linear detection range of 500 fg mL-1 to 100 ng mL-1 under optimal conditions. The proposed immunosensor also exhibits outstanding selectivity, storage stability, reproducibility, and regeneration capabilities. Application of the immunosensor to clinical samples (Positive: n=10; healthy: n=5) demonstrated high accuracy, with results aligning well with those from the enzyme-linked immunosorbent assay (ELISA) technique. The findings showed that the suggested sensing platform could be utilized for the early diagnosis of cervical cancer.