Appl

Appl. planes of the AuNPs, respectively (curve b) 38. 3.3. Electrochemical behavior of the biosensor The assembly processes of the altered electrode were verified by cyclic voltammetry (CV) and electrochemical impedance spectroscopy techniques (Physique?2). As can be seen in Physique?2A, the CV curve of GC electrode in Biotin-X-NHS [Fe (CN)6]4? media (curve a) have a redox peak related to Fe3+/2+ electrochemical reaction. After modification of GCE surface with GO/AuNPs (curve b), the electrochemical active surface area has Biotin-X-NHS significantly enhanced, indicating that GO/AuNPs increases the active surface of electrode for the conjugation of biomarker. Immobilization of antibodies and antigen on the surface of electrode (curves cCe) Biotin-X-NHS leads to a decrease in the electrochemical active surface area of electrode due to the hindering effect on the electron transfer rate that results in the attenuation of redox peaks 39. Physique?2B shows the Nyquist plots of sensing electrode confirming and the step\by\step modification of the electrode according to the procedure mentioned in Experimental Section and Scheme?1. Open in a separate Biotin-X-NHS window Physique 2 (A) Electrochemical CV responses of bare GC electrode (a),G/GNP (b) G/GNP/Ab1 (c), G/GNP/Ab1/Ag(d), and G/ GNP/Ab1/Ag/Ab2 (e). (B) Electrochemical impedance spectra of bare GC electrode (a), G/GNP (b), G/GNP/Ab1 (c), G/GNP/Ab1/Ag(d), and G/ GNP/Ab1/Ag/Ab2 (e), (C) Effects of the incubation time. (D) And volume for GO/ AuNPs /Ab2 nanoprobes Biotin-X-NHS around the ip of the sensor. All measurements were recorded in 0.1?M KCl solution containing 4?mM [Fe(CN)6]4?/3? As presented, each plot possesses a semicircle that is ascribed to charge transfer resistance for electrochemical reaction of Fe3+/2+ ions on the surface of electrode. A minuscule semicircle domain name for the bare GC electrode was observed (curve a), implying a fast electron transfer process. Subsequently, GO/AuNPs were deposited on the surface of GCE. As shown, the diameter of the semicircle decreased (curve b) due to increasing electrochemical active surface area of the electrode that facilitates electron transfer. After immobilization of GO/AuNPs /Ab1, a conspicuous escalation of the semicircle diameter was also observed (curve c), implying a apparent augmentation of electrochemical impedance. Moreover, an apparent increase of the resistance with sequential assembly of antigen PSA can be discerned (curve d). It is also obvious that this addition of GO/AuNPs/Ab2 caused a gradual increase in the resistance of the electrode as the experiment proceeded (curve e). These results confirmed that this antibodyCantigen nanoprobe sandwich\like architecture has been successfully assembled. The effect of incubation time and volume of nanoprobe answer on detection of total PSA are studied (Physique?2C and D). As shown, a minor increase in the incubation time leads to a significant decrease of ip. The results show that after 20?min, the response barely changes (about 32%). Therefore, the incubation time for the antigen and antibody incubation was found to be about 20 min. In addition to above mentioned results, the impact of the volume of nanoprobe answer was further investigated. Physique?2D presents the relationship between ip and the volume of nanoprobe answer. Therefore, the intensity of peak current density linearly increases with an increase in the volume of the nanoprobe answer until it reaches to 20 L. In continuation, from 20 to 60 L, the plot reveals that this intensity of the peak current density is usually approximately independent of the volume of nanoprobe answer. These results indicate that this optimum volume answer for the nanoprobe is about 20 L. 3.4. Detection of the total and free PSA The responses of square wave voltammetry of total PSA and free PSA are illustrated in Physique?3A and B. Physique?3A shows that SW (square wave voltammetry) responses decrease with increasing concentration of total PSA. Furthermore, square Rabbit Polyclonal to RAB5C wave voltammetry (SW) responses decrease with increasing concentration of Free PSA (3B). LOD is usually defined as the lowest possible concentration of an analyte that can be distinguished without any assurance about the imprecision of the obtained results. Herein, the LOD was calculated by the following equation 37: LOD is the sensitivity of the sensor (slope of the regression line) and is the SD of the blank. Mean value and SD of ip were calculated, since each concentration measurement was repeated three times (Physique?3C and D). According to Equation?(1), the LOD of total PSA and free.