Biosensors

Aptamer based biosensors (Aptasensors) are well-constructed by a variety of methodologies, including electrochemical biosensors, optical biosensors and mass-sensitive biosensors. The design strategies of most biosensors have some similar elements. These strategies can be divided into four modes:

1. Target-induced structure switching mode (TISS)

Binding of the targets to the immobilised aptamer causes the aptamers to alter configuration from a flexible to rigid tertiary structure such as G-quaduplex. Such conformational switches would change the relative positions of signal moieties, leading to initiation of a signal.Examples of biosensors using TISS include  Aptamer Beacons .

A reported example features methylene blue incorporation into the aptamer stem-loop structure (Fig. 1). Upon target binding, methylene blue is separated from the surface resulting in a change in signal.

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Figure 1. Design of a signal-off electrochemical aptamer biosensor for thrombin 1.

2. Sandwich like mode

An aptamer is immobilized onto a surface as a capture probe and a second aptamer is modified with a label as a signal read-out probe. The target can be captured on the surface of electrodes by binding to the capture aptamer and then forming a sandwich complex with the second aptamer. During this process, sandwiched binding complexes bring catalytic labels to the surfaces of electrodes to generate colorimetric, voltammetric, impedimetric, or gravimetric signals for detection.

A reported example was an electrochemical biosensor for Platelet-derived growth factor (PDGF) detection.  The “sandwich” structure is fabricated based on the fact that PDGF has two aptamer-binding sites, which makes it possible for one PDGF molecule to connect with two aptamers simultaneously. Gold nanoparticles (Au-NPs) Allowed to amplify the signal of the electrochemical probe [Ru(NH3)5Cl](2+) to obtain a very low detection limit for the analysis of PDGF in serum samples (Fig.2).

Figure 2. Schematic representation of the electrochemical aptasensor based on a sandwich assay and on the use of [Ru(NH3)5Cl](2+) as a redox probe.

Figure 2. Schematic representation of the electrochemical aptasensor based on a sandwich assay and on the use of [Ru(NH3)5Cl](2+) as a redox probe (2).

 3.Target-induced dissociation mode (TID)

In TID mode, the complementary sequences of aptamers, instead of aptamers themselves, are employed as anchors to localize the aptamers. After incubation with targets, the formed target-aptamer complexes will be liberated into the solution, which leads to the changes of detectable signals. TID strategy can be further classified into signal-off mode, signal-on mode, and label-free mode, as shown in figure 3.

Figure 3. Schemes for the target-induced dissociation/displacement strategies. A. signal off mode ; B signal-on mode ; C label-free mode.

Figure 3. Schemes for the target-induced dissociation/displacement strategies. A. signal off mode ; B signal-on mode ; C label-free mode (3).

4. Competitive replacement mode

In a competitive assay, One component (either the aptamer or the analyte) is immobilised on the sensor’s surface. In the case of immobilised Aptamers a label target is bound to the aptamer. Competitive replacement of the labelled target by an unlabelled target originating from the sample results in a decrease of the signal. In this mode, signal modified target molecules need to be designed and synthesized in advance.

A reported example is the development of an ultra-sensitive electrochemical biosensor for the detection of thrombin and lysozyme simultaneously.The thiolated aptamers of thrombin and lysozyme are co-immobilized onto the gold substrate, followed with the binding of cadmium sulphide (CdS)-labeled thrombin and lead sulphide (PdS)-labeled lysozyme. As the sample containing detection targets was added, the label-proteins could be displaced by targets, which could be characterized through electrochemical stripping detection (Fig 4).

Figure 4. Dual-analyte biosensor designed via the competitive replacement mode and electrochemical stripping detection (4).

Figure 4. Dual-analyte biosensor designed via the competitive replacement mode and electrochemical stripping detection (4).

Nanoparticles

Conjugation of aptamers on various nanomaterials has led to highly sensitive and selective aptasensors.Examples of nanomaterials used for biosensors include metallic nanoparticles, quantum dots, silica nanoparticles, and carbon nanotubes, with interest in aptamer incorporation increasing in popularity.

 

We are currently collaborating with several parties to develop aptamer based biosensors for the reliable detection of disease associated biomarkers or field based contaminants/pathogens in a decentralised setting. We see the successful integration of aptamers as a key step toward solving the increasing need for highly sensitive, rapid and economical diagnostic devices.