Quantification of Tumor Protein Biomarkers from Lung Patient Serum Using Nanoimpact Electrochemistry

Abstract

Protein quantification with high throughput and high sensitivity is essential in the early diagnosis and elucidation of molecular mechanisms for many diseases. Conventional approaches for protein assay often suffer from high costs, long analysis time, and insufficient sensitivity. The recently emerged nanoimpact electrochemistry (NIE), as a contrast, allows in situ detection of analytes one at a time with simplicity, fast response, high throughput, and the potential of reducing the detection limits down to the single entity level. Herein, we propose a NIE-enabled electrochemical immunoassay using silver nanoparticles (AgNPs) as labels for the detection of CYFRA21-1, a typical protein marker for lung carcinoma. This strategy is based on the measurement of the impact frequency and the charge intensity of the electrochemical oxidation of individual AgNPs before and after they are modified with anti-CYFRA21-1 and in turn immunocomplexed with CYFRA21-1. Both the frequency and intensity modes of single-nanoparticle electrochemistry correlate well with each other, resulting in a self-validated immunoassay that provides linear ranges of two orders of magnitude and a limit of detection of 0.1 ng/mL for CYFRA21-1 analysis. The proposed immunoassay also exhibits excellent specificity when challenged with other possible interfering proteins. In addition, the CYFRA21-1 content is validated by a conventional, well-known enzyme-linked immunosorbent assay and successfully quantified in a diluted healthy serum with a satisfactory recovery. Moreover, CYFRA21-1 detection in serum samples of lung cancer patients is successfully demonstrated, suggesting the feasibility of the NIE-based immunoassay in clinically relevant diagnosis. To the best of our knowledge, this is the first report to construct NIE-based electrochemical immunoassays for the specific detection of tumor protein biomarkers.

From Nanoparticle Ensembles to Single Nanoparticles: Techniques for the Investigation of Plasmon Enhanced Electrochemistry

“…NIE, different from the measurement of single immobilized nanoparticles, is a technique based on the stochastic collisions of individual random-walk nanoparticles in solution onto an inert UME. [36][37][38][39] In a typical NIE mode, the electrochemically active molecules in solution react on the surface of the individual nanoparticles when they collide with the UME, resulting in transient current spikes appearing on a current-time profile. By controlling the concentration of the nanoparticles in a certain range, one spike current can be corresponded to one event of a single nanoparticle collision, thus resulting in a single nanoparticle sensitivity.…”

“…PEEC at a real “single nanoparticle” level is yet very rare to see using a conventional electrochemical workstation due to the high requirement of the detection sensitivity and the operational complexity of positioning a single nanoparticle on a micro/nano substrate. NIE, different from the measurement of single immobilized nanoparticles, is a technique based on the stochastic collisions of individual random‐walk nanoparticles in solution onto an inert UME [36–39] . In a typical NIE mode, the electrochemically active molecules in solution react on the surface of the individual nanoparticles when they collide with the UME, resulting in transient current spikes appearing on a current‐time profile.…”

A general controllable release amplification strategy of liposomes for single-particle collision electrochemical biosensing

“…The application of SPCE in biochemical analysis has attracted wide attention credited to its ultrasensitivity up to single-particle level and reliable quantitative model. Which can be mainly divided into two categories: 1) The direct collision of biological macromolecules or insulating particles with a microelectrode, catalyzes or hinders oxidation-reduction active molecules ( Dick et al, 2015 , 2016 ; Zhang et al, 2021 ). This method is simple, fast, and label-free, but lacks selectivity and specificity, in addition to the interference of complex system has non-negligible influence.…”

Single DNA Origami Detection by Nanoimpact Electrochemistry

“…[4] This method has been used with inorganic and organic nanoparticles, [5] emulsion droplets, [6] lipid vesicles, [7] bacteria, [8] viruses and enzymes, [9] . It has also been utilised as a biosensor to indirectly detect tumour biomarkers [10] and viral DNA. [11] However, so far, little work has been focused on the detection of DNA self-assembled nanostructures.…”