**2. Electrochemical Biosensors**

Electrochemistry is essential for achieving the biosensing process in various biomarker analyses. Thus, electrochemical biosensing has attracted widespread attention in various applications due to its considerable advantages. Electrochemical biosensors react with the analyte of interest or molecules to produce an electrical signal proportionate to the analyte concentration. A conventional electrochemical biosensor comprises a reference electrode and a sensing electrode (working electrode) separated by an electrolyte. In most applications, the electrochemical biosensors consist of a three-electrode system with the reference electrode connected to a potentiostat, and the circuit can be completed by adding a counter electrode for flowing the current. These sensing devices are inexpensive, low-cost electrochemical cells that can be produced, portable, and easy to use, and can be operated with reduced power consumption. It requires electronic components for detecting the target analytes, unlike optical sensors. The following sections describe a range of elements and techniques of the electrochemical biosensor for biosensing applications.

#### **3. Surface Modification Technique**

Surface chemistry plays a considerable role in electrochemical biosensors to link the biological recognition element (BRE) on top of the sensing surface and prevent the substrate electrode from nonspecific interactions. In addition, the functionalization of the surface is conducive to noise control and sensitivity enhancement. BRE used in electrochemical sensors mainly consists of enzymes, antibodies, DNA/RNA, aptamers, and whole cells [22], which define a biosensor's sensitivity and selectivity. Immobilization techniques, such as adsorption, encapsulation in polymers or gel, chemical crosslinking, self-assembled monolayer, covalent linking, affinity, and electrodeposition, have been widely investigated for detecting various analysts in complex water samples. The surface modification of BRE on the electrodes usually involves one or more strategies.

## *3.1. Adsorption*

Adsorption is a straightforward method of modifying the surface of the electrode to a specific recognition element in an entirely arbitrary way. Every biological recognition element needs to achieve the best conditions. Most proteins usually achieve the best surface coverage on uncharged surfaces under the neutral pH and functional ionic strength, using a specific 5−20 μg/mL concentration [35]. Yang et al. [36] have developed an impedimetric-based immunosensor by the adsorption method of anti-*E. coli* antibodies against the integrated microelectrode arrays for detecting the *E. coli* O157:H7. Surface modification of the surface receptor proteins G and A can be produced by many bacterial strains that can promote receptor binding. Each protein, such as A and G, can certainly be capable of binding 4 and 2 molecules of IgG. An analogous method utilizes or takes advantage of the strong binding of the glycoprotein avidin for biotin-functionalizing the receptors due to the intense alienation of the avidin/biotin complex. The detection level of *E. coli* was 1.3 × <sup>10</sup>−<sup>15</sup> M, which was as low as 10−100 CFU mL−<sup>1</sup> in concentration. It may be identified on the avidin-modified developed electrodes using biotinylated anti-*E. coli* as the targeted recognition ligand.
