Electrochemical Genosensing of Circulating Biomarkers

Abstract

Management and prognosis of diseases requires the measurement in non- or minimally invasively collected samples of specific circulating biomarkers, consisting of any measurable or observable factors in patients that indicate normal or disease-related biological processes or responses to therapy. Therefore, on-site, fast and accurate determination of these low abundance circulating biomarkers in scarcely treated body fluids is of great interest for health monitoring and biological applications. In this field, electrochemical DNA sensors (or genosensors) have demonstrated to be interesting alternatives to more complex conventional strategies. Currently, electrochemical genosensors are considered very promising analytical tools for this purpose due to their fast response, low cost, high sensitivity, compatibility with microfabrication technology and simple operation mode which makes them compatible with point-of-care (POC) testing. In this review, the relevance and current challenges of the determination of circulating biomarkers related to relevant diseases (cancer, bacterial and viral infections and neurodegenerative diseases) are briefly discussed. An overview of the electrochemical nucleic acid-based strategies developed in the last five years for this purpose is given to show to both familiar and non-expert readers the great potential of these methodologies for circulating biomarker determination. After highlighting the main features of the reported electrochemical genosensing strategies through the critical discussion of selected examples, a conclusions section points out the still existing challenges and future directions in this field.

Keywords: bacterial and viral infections; cancer; circulating biomarkers; electrochemical genosensors; neurodegenerative diseases.

Figure 1

Schematic display of a general nucleic acid-based sensor for the determination of a particular target involving immobilization of a specific capture probe on the electrode surface (1); specific hybridization of the immobilized capture probe with the target sequence (2) and of the captured target with the label-conjugated detector probe (3) and electrochemical detection of the hybridization reactions by differential pulse voltammetry of the label attached to the detector probe (4).

Figure 2

Schematic display of the target-responsive encapsulation nucleic acid-based electrochemical biosensor functioning and the DPV signals obtained in the presence and in the absence of the target DNA. Reprinted and adapted from [48] with permission.

Figure 3

Electrochemical biosensor for HIV antibodies using a specific peptide and short DNAs as diluents. In the presence pf HIV antibodies the surface mobility of the immobilized peptide probe is limited and the MB current measured by ACV decreased. Reprinted from [50] with permission.

Figure 4

Schematic display of the MBs-based amperometric DNA sensor developed for S. pneumoniae determination through the detection of the ss-aPCR amplicon generated from a specific fragment of lytA gene coding sequence by performing daPCR directly in bacterial cultures. Reprinted and adapted from [55] with permission.

Figure 5

Schematic display of the electrochemical DNA biosensor developed for Mycobacterium sp. genomic DNA detection using a specific probe immobilized on an ITO electrode and AuNPs modified both with detector probes and the enzyme alkaline phosphatase (AP). Reprinted from [58] with permission.

Figure 6

Schematic representation of the steps involved in the M. tuberculosis determination using an electrochemical genomagnetic assay coupled to HDA. The biotinylated ss-amplicon resulting from the HDA was immobilized onto the surface of Strep-MBs and further hybridized with an FITC-detector probe and labeled with an HRP-antiFITC antibody. Reprinted from [62] with permission.

Figure 7

Schematic representation of the impedimetric genosensor developed for L. infantum determination. Reprinted from [65] with permission.

Figure 8

Scheme of the electrochemical method developed to determine Leishmania DNA by using RPA and primers labeled with AuNPs and MBs involving the following steps: DNA extracted from dog blood (a), isothermal amplification of a specific region by RPA using primers labeled with AuNPs and MBs (b), magnetic capture of the MB/amplified DNA/AuNP complexes of the SPCE (c) and chronoamperometric detection of the hydrogen evolution reaction (HER) by AuNPs (d). Reprinted from [66] with permission.