# Recent Advances in Electrochemical Immunosensors
## 3 Discussion
### Metal or Metal Oxide Nanoparticles
#### 3.4.2 Enzyme-Free Immunosensors
With a sandwich-type immunosensor, four tumor markers of pancreatic cancer (CEA, CA199, CA125 and CA242) were detected. The reported linear response ranges were from 0.1 to 100 ng mL\({}^{-1}\) for CEA and 1 to 150 U mL\({}^{-1}\) for CA199, CA125 and CA242. LoDs were of 0.02 ng mL\({}^{-1}\), 0.4 U mL\({}^{-1}\), 0.3 U mL\({}^{-1}\) and 0.4 U mL\({}^{-1}\), respectively (1 enzyme unit (U) = 1 \(\upmu\)mol min\({}^{-1}\)). Also for measuring CEA, [PERSON] et al. [134] reported a thionine/infinite coordination polymer functionalized by AgNPs used to increase the overall area as well as the electron transfer rate. Using DPV and Fe(CN)\({}_{6}\)\({}^{3-/4-}\) in solution, the authors found a linear range between 50 fg mL\({}^{-1}\) and 100 ng mL\({}^{-1}\) and a LoD of 0.5 fg mL\({}^{-1}\). These good results underline the interest of infinite coordination polymer particles. They are of great interest because these structures present a high level of structural tailorability, which allows to tune microporosity and catalytic activity. Still for CEA detection and using Fe(CN)\({}_{6}\)\({}^{3-/4-}\) as redox probe in solution, [PERSON] et al. [135] described a AuNPs-modified Au electrode onto which protein A was adsorbed for oriented immobilization of anti-CEA antibodies. Using EIS and CV, they obtained a linear response to CEA between 1 pg mL\({}^{-1}\) and 100 ng mL\({}^{-1}\), and a LoD of 0.1 pg mL\({}^{-1}\). [PERSON] et al. [136] reported an enzyme-free paper-based electrochemical immunosensor for detection of CEA. Amino-functionalized graphene/thionine/AuNPs were synthesized and deposited on the electrode along with CEA antibodies. The decreased CV current of Thi was proportional to the concentrations of corresponding antigens within the range 50 pg mL\({}^{-1}\)-500 ng mL\({}^{-1}\), with a LoD of  
Figure 23: **(A)**_P/_HRP2 immunosensor in the absence **(a)** and presence **(b)** of the target antigen with the MB-labelled detection antibody; **(B)** Chronocoulograms of PBS spiked with _P/_HRP2 recorded at 0.05 V (vs. Ag/AgCl). Reprinted from [122], Copyright 2016, with permission from Elsevier; **(C)** Scheme of **(a)** construction of signaling AuNPs by layer-by-layer alternate assembly of PDDA and PSS, and **(b)** sandwich immunoassay with Ag deposition and AgNPs redissolution, for CEA detection, on a chitosan/RGO-modified GC electrode; **(D)** Linear sweep stripping voltammetric curves of AgNPs for detection of CEA from 0 to 0.5 ng mL\({}^{-1}\). Reprinted with permission from [127]. Copyright 2012 American Chemical Society. 10 pg mL\({}^{-1}\). [PERSON] et al. [137] used impedance on interdigitated electrodes (IDEs) to detect CEA with PANI-AuNPs as labels. In a first step, adamatly-modified capture antibodies were immobilized on IDEs functionalized with carboxymethyl-\(\beta\)-cyclodextrintrim. After immunoreaction between the capture antibodies and CEA, a PANNi-AuNPs-labeled signaling antibodies were added. This recognition sequence led to a linear resistance decrease from 0.1 ng mL\({}^{-1}\) to 1000 ng mL\({}^{-1}\) and a LoD of 7 pg mL\({}^{-1}\). [PERSON] et al. [138] reported the use of AuNPs on thionine-modified MoS\({}_{2}\) nanosheets for electrochemical label-free immunosensing of CEA, where Thi was used not only as a redox indicator but also as a reducing agent for synthesis of the AuNPs. Several shapes of AuNPs were investigated from spherical, triangle, clover-like to flower-like shapes. This system could detect CEA with a LoD of 0.5 pg mL\({}^{-1}\). At last, also for CEA detection, [PERSON] et al. [139] demonstrated Ag/AuNPs/GR used as both substrate and label of signaling antibodies. Using a sandwich-type immunoassay and CV to measure oxidation/reduction of the Ag\({}^{0}\)/Ag\({}_{2}\)O couple (Figure 24C), the authors reported a linear calibration range from 10 pg mL\({}^{-1}\) to 1.2 \(\times\) 10\({}^{5}\) pg mL\({}^{-1}\), and a LoD of 8 pg mL\({}^{-1}\). Detection of other classical proteins were reported, such as HSA (a model protein) and PSA. [PERSON] et al.