Study on a new high affinity anti-glypicans-3 antibody in diagnosis of early hepatocellular carcinoma by differential pulse voltammetry

A novel electrochemical immunoassay for glypicans-3 (GPC3) was established based on its new high affinity antibody (Ig). In this study, we used glass carbon film (GCF) as carrier for immobilizing Ig and avoiding the high electron transfer resistance (Ret) in electrochemical immunoassay sensor (Ig-Biotin-GR/GCE) because many materials such as bovine serum albumin (BSA), avidin, biotin, Ig, GPC3, and ALP were covered on. The Ig was combined with GCP3 labeled alkaline phosphatase (ALP-GPC3) which catalyzes p-nitro phenyl phosphate (PNPP) into p-nitro phenol (PNP). In order to obtain highly sensitive electrochemical signals of the PNP and further determine ALP-GPC3, the GR/GCE modified polylysine (PLYS/GR/GCE) was fabricated and used for detecting the PNP. According to the principle of competitive inhibition, less ALP-GPC3 would be combined with the Ig in the presence of GPC3. By investigating the signals of PNP in the absence and presence of GPC3, GPC3 could be qualitatively and quantitatively analyzed. Under the optimal conditions, the peak current of PNP on the PLYS/GR/GCE was linear with the concentration of GPC3 in the range of 0.7575 ng mL(-1) and a detection limit of 0.2 ng mL(-1). The recognition of GPC3 and detection of PNP were separated, which provides a new way for studying the diagnosis of liver cancer.