# Recent Advances in Electrochemical Immunosensors
## 3 Discussion
### Diffusing Redox Probes
#### 3.3.2 Enzyme-Based Immunosensors
\begin{table}
\begin{tabular}{c c c c} \hline \hline
**Target** & **Probe** & **LoD** & **Method** & **Reference** \\ \hline PSA & Ab & 13 pg mL\({}^{-1}\) & Graphene/MB/CV & [70] \\ Kanamycin & Ab & 6 pg mL\({}^{-1}\) & GR/PtNPs/Thi/CV & [71] \\ Kanamycin & Ab & 15 pg mL\({}^{-1}\) & Ag/Fe\({}_{3}\)O\({}_{4}\) NPs/Thi/SWV & [72] \\ CEA protein & Ab & 5 pg mL\({}^{-1}\) & PdCu-GR/Thi/CV & [73] \\ CEA protein & Ab & 10 pg mL\({}^{-1}\) & Carbon CMK-3/Thi/DPV & [74] \\ CEA protein & Ab & 0.05 fg mL\({}^{-1}\) & AuNPs/Thi & [75] \\ CEA protein & Ab & 0.024 pg mL\({}^{-1}\) & Ag/NPs/ASV & [76] \\ CEA protein & Ab & 0.1 ng mL\({}^{-1}\) & GR/Toluiudine blue & [77] \\ CEA protein & Ab & 3.3 fg mL\({}^{-1}\) & AuNPs/Toluiudine blue/DPV & [78] \\ CEA protein & Ab & 0.4 pg mL\({}^{-1}\) & GR/CHI/Fc & [79] \\ CEA protein & Ab & 0.5 pg mL\({}^{-1}\) & GO/AuNPs/NB & [80] \\ AFP protein & Ab & 5 pg mL\({}^{-1}\) & PdNPs/RGO/Amprometry & [81] \\ AFP protein & Ab & 0.1 fg mL\({}^{-1}\) & Cu\({}_{2}\)O/GO/TB/SWV & [82] \\ Avian leukosis virus & Ab & 115 TICID\({}_{50}\) mL\({}^{-1}\) & Fe\({}_{3}\)O/CuNPs/ASV & [83] \\ Tumor cells & Ab & 10 cells mL\({}^{-1}\) & CdTe/SWV & [84] \\ Interleukin-6 & Ab & 5 fg mL\({}^{-1}\) & GR nanoribbon/AgNPs & [85] \\ Estradiol & Ab & 0.015 pg mL\({}^{-1}\) & Fe\({}_{3}\)O\({}_{4}\)/Pb\({}^{2+}\)/ASV & [86] \\ Cardiac troponin-I & Ab & 0.05 ng mL\({}^{-1}\) & GO/AuNPs/Fc/SWV & [87] \\ Myoglobin & Ab & 0.01 ng mL\({}^{-1}\) & Graphene QDs/EIS & [88] \\ miRNA & Ab & 5 fmol L\({}^{-1}\) & RGO/PCE/SWV & [89] \\ \hline \hline \end{tabular}
\end{table} 
Table 5: Figures of merit of selected enzyme-free immunosensors using graphene and immobilized redox probe. polydopamine as linker. HRP-labeled capture anti-CEA were immobilized on the lectin through the specific sugar-protein interaction (specific affinity between concanavalin A and sugar chains located on the surface of HRP). With Fe(CN)\({}_{6}\)\({}^{3-/4-}\) as redox probe and DPV for measurements, CEA was detected within a linear range of 0.1-750 ng mL\({}^{-1}\) (LoD of ca. 90 pg mL\({}^{-1}\)). This approach must attract our attention because, conversely to conventional grafting of antibodies on a solid substrate, which cannot be well controlled in terms of Ab orientation on the surface, this immobilization by affinity insures that all probe antibodies are well oriented on the surface and are available for further target (antigen) recognition. Still for CEA detection, [PERSON] et al. [95] described a streptavidin-functionalized nitrogen-doped graphene where biotinylated antibodies were immobilized. HRP-labelled antibodies were used with a sandwich immunoassay, with Thi and H\({}_{2}\)O\({}_{2}\) as redox probes. With DPV, the authors demonstrated a linear detection range from 0.02 to 12 ng mL\({}^{-1}\) and a LoD of 0.01 ng mL\({}^{-1}\). Graphene-based immunosensors were also designed for detection of hormones. For example, [PERSON] et al. [96] described in 2013 a 17\(\beta\)-estradiol immunosensor based on a GR/PANi composite as electrode substrate, and HRP-labelled signaling antibodies coupled to GO sheets. They used a competitive immunoassay with HQ as redox mediator. With DPV, they obtained a linear response to estradiol in the range 0.04-7.00 ng mL\({}^{-1}\) and a LoD of 0.02 ng mL\({}^{-1}\). [PERSON] et al. [97] also reported ethynylestradioliol (EE2) detection using a AgNPs/SiO\({}_{2}\)/GO composite grafted on a GC electrode through the diazonium route, onto which anti-EE2 was also covalently bound. HRP-EE2 was used in a competitive assay, with HQ as substrate. The current was monitored at a constant potential of \(-\)200 mV and led to a LoD of 65 pg mL\({}^{-1}\) and a linear range between 0.1 and 50 ng mL\({}^{-1}\). [PERSON] et al. [98] reported very recently (2017) detection of cortisol using AuNPs/GO. Cortisol, was first coupled to AuNPs/GO immobilized on a GCE by a Nafion membrane.