Bio-assay of the non-amidated progastrin-derived peptide (G17-Gly) using the tailor-made recombinant antibody fragment and phage display method: a biomedical analysis

Abstract

In this research, four novel and sensitive immunosensors for electrochemical determination of G17-Gly were designed based on signal amplification and tailor-made recombinant antibody technology. Anti-G17-Gly antibody fragments (i.e. scFv and VL specific to the N- and C-terminal of G17-Gly) were immobilized onto a polymeric nanocomposite comprising poly cetyl trimethyl ammonium bromide (P(CTAB)) as the conductive matrix, chitosan (CS) as a biocompatible agent and gold nanoparticles (AuNPs) as the signal amplification element. The high surface area provided by AuNPs and the small size of scFv/VL establish the basis for immobilizing a high amount of the anti-G17-Gly on the surface of the electrode for detecting G17-Gly in human plasma samples. Under optimal conditions, the designed immunosensors provide an excellent analytical capability for detecting and determining G17-Gly in human plasma samples with a linear range from 0.5 mM to 0.05 pM and a LLOQ of 0.05 pM. The sensitivity order of the immunosensors was Ag/2-mercaptoethanol/phage displaying scFv/P(CTAB–CS)–AuNP/GE, Ag/2-mercaptoethanol/phage displaying VL/P(CTAB–CS)–AuNP/GE, Ag/BSA/scFv/P(CTAB–CS)–AuNP/GE, and Ag/BSA/VL/P(CTAB–CS)–AuNP/GE. The aforementioned characteristics demonstrate that the proposed immune-devices can be used in biological and clinical diagnosis as reliable tools for identifying different oncobiomarkers.