Ultrasensitive bioassaying of HER-2 protein for diagnosis of breast cancer using reduced graphene oxide/chitosan as nanobiocompatible platform

Hassan Nasrollahpour,Ibrahim Isildak,Balal Khalilzadeh,Abdolhosein Naseri,Mohammad-Reza Rashidi,Esmat Alsadat Hashemi

doi:10.1186/s12645-021-00082-y

Abstract

BackgroundIn this label-free bioassay, an electrochemiluminescence (ECL) immunosensor was developed for the quantification of breast cancer using HER-2 protein as a metastatic biomarker.MethodFor this purpose, the ECL emitter, [Ru(bpy)3]2+, was embedded into biocompatible chitosan (CS) polymer. The prepared bio-composite offered high ECL reading due to the depletion of human epidermal growth factor receptor 2 (HER-2) protein. Reduced graphene oxide (rGO) was used as substrate to increase signal stability and achieve greater sensitivity. For this, rGO was initially placed electrochemically on the glassy carbon electrode (GCE) surface by cyclic voltammetry (CV) technique. Next, the prepared CS/[Ru(bpy)3]2+ biopolymer solution was coated on a drop of the modified electrode such that the amine groups of CS and the carboxylic groups of rGO could covalently interact. Using EDC/NHS chemistry, monoclonal antibodies (Abs) of HER-2 were linked to CS/[Ru(bpy)3]2+/rGO/GCE via amide bonds between the carboxylic groups of Ab molecules and amine groups of CS. The electrochemical behavior of the electrode was studied using different electrochemical techniques such as electrochemical impedance spectroscopy (EIS), differential pulse voltammetry (DPV) and square wave voltammetry (SWV) and also ECL tests.ResultsAfter passing all optimization steps, the lower limit of detection (LLOQ) and linear dynamic range (LDR) of HER-2 protein were practically obtained as 1 fM and 1 fM to 1 nM, individually. Importantly, the within and between laboratory precisions were performed and the suitable relative standard deviations (RSDs) were recorded as 3.1 and 3.5%, respectively.ConclusionsAs a proof of concept, the designed immunosensor was desirably applied for the quantification of HER-2 protein in breast cancer suffering patients. As a result, the designed ECL-based immunosensor has the capability of being used as a conventional test method in biomedical laboratories for early detection of HER-2 protein in biological fluids.Graphic

Highlights

In this label-free bioassay, an electrochemiluminescence (ECL) immu‐ nosensor was developed for the quantification of breast cancer using human epidermal growth factor receptor 2 (HER-2) protein as a metastatic biomarker
After passing all optimization steps, the lower limit of detection (LLOQ) and linear dynamic range (LDR) of HER-2 protein were practically obtained as 1 fM and 1 fM to 1 nM, individually
As a proof of concept, the designed immunosensor was desirably applied for the quantification of HER-2 protein in breast cancer suffering patients

Summary

Introduction

In this label-free bioassay, an electrochemiluminescence (ECL) immu‐ nosensor was developed for the quantification of breast cancer using HER-2 protein as a metastatic biomarker. Global concerns of BC spreading, high prices for curing and diagnostic drawbacks triggering the demands for costly effective, rapid and on-site recognition of this disease (Min et al 2018; Huang et al 2019; Zamani-Ahmadmahmudi et al 2014). Among many advancements in bioanalytical domain, emerging biosensors regarded as a milestone for future hopes. These devices have been trusted by many diagnostic domains for recognition of diseases. [Ru(bpy)3]2+ has attracted great success for in vitro recognition purposes. This is due to its superb sensitivity, easy to use and broad dynamic range. Coreactant-assisted ECLs of ruthenium complexes (including [Ru(bpy)3]2+) play substantial act in analytical and bioanalytical applications. Tri-propyl amine (TPrA) is a well-known coreactant of [Ru(bpy)3]2+ which is widely employed in laboratorial and commercial assays (Richter 2004; Liu et al 2015; Li et al 2017; Zanut et al 2020)

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