Probe Functionalization With A Rhop-3 Antibody: Toward A Rhop-3 Antigen Immunosensor For Detection of Malaria

The antibody specific for the malaria protein, Rhop-3, and FL-Rhop-3, were immobilized on the surface of a gold electrode modified with cysteamine. Colloidal gold was used to enhance the detection signal for Rhop-3 antigens. The Rhop-3 antibody was also immobilized on gold electrodes preactivated with dithiobis(succinimidyl proprionate) (DSP). Immobilization was performed at room temperature and at 37 °C. Cyclic voltammetry (CV) was used to monitor the interaction between the immobilized antibody and its cognate antigen in solution, using ferricyanide, K3Fe(CN)6, as reporting electroactive probe. Tests indicate recognition of Rhop-3 protein by the immobilized antibody. Antigen recognition was enhanced by incubation at 37 °C compared with room-temperature incubation. Our results suggest that an immunosensor can be developed and optimized to aid detection of Rhop-3 antigens in samples from malaria patients. As far as we are aware, this is the first amperometric immunosensor targeting Rhop-3 antigen as a malaria biomarker

Zinc Metal at a New ZnO Nanoparticles Modified Carbon Paste Electrode: A Cyclic Voltammetric Study

A new carbon paste electrode modified with ZnO nanoparticles was prepared and used to study the electro oxidation of zinc ions in solution by cyclic voltammetry (CV) method. The modified electrode displayed strong resolving function for the overlapping voltammetric response of zinc into one well-defined peak. The potential difference between Epa and Epc was > 200 mV, the range of which referred to the quasi-reversible mechanism. The kinetic of electrode was studied in the range of temperature 15-30 ℃. The data of voltamograms showed the negative shift increased with increase of temperature, the result of which referred to the diffusion electron transfer in redox process of zinc oxidation. Diffusion coffining was calculated from Sevcik-Randles equation and thus equaled to 1.01×10-7. The rate constant was equal to 5.2×10-8 and 3.3×10-8 for oxidation and redaction respectively. And the peak current of zinc increased linearly with its concentration in the range of 2.0-20.0 ppm

Design of Ultrasensitive Electrochemical Aptasensor for Earlier Detection of Hepatitis B Virus

In consideration of the imperious need for developing a simple, ultra-sensitive and selective method to detect Hepatitis B virus (HBV), an label free aptasensor was constructed by the modification of a chemically inert glassy carbon electrode (GCE) by dint of a gold nanoparticle immobilized reduced graphene oxide. The ultrasensitive detection was attained using a specific ssDNA (thiolated aptamer at the 5′ end) as a recognition element. The aptamer was fastened onto GCE/rGO/Au surface via the Au-S bond for determining the impedance of charge mobility by means of [Fe(CN)6]3-/4- as a redox gauge. The function of aptasensor based on the specific interaction between the aptamer and specific surface antigen HBsAg induced [HBsAg-aptamer] complex formation which impeded electron movability between a [Fe(CN)6]3-/4- redox probe solution and sensing interface. HBsAg-binding aptamer events can be easily monitored by the signal switch through cyclic voltammetry technique (CV), square wave voltammetry (SWV), and electrochemical impedance spectroscopy (EIS) measurements. Under the optimized conditions, the aptasensor can detect HBsAg in the 0.25 fg/mL to 1.5 fg/mL concentration range and limit of detection of 0.018, 0.0016 and 0.001 fg/mL for CV, SWV and EIS respectively. The recovery of 88-109.6% was obtained. Moreover, the selectivity of the aptasensor was tested using other biological interfering substances. This developed aptasensor provided one of the lowest limits of detection for HBsAg recently reported in the literature, satisfying stability and good reproducibility, consequently could be applied for revealing HBsAg in real samples

Probe Functionalization With A Rhop-3 Antibody: Toward A Rhop-3 Antigen Immunosensor For Detection of Malaria

The antibody specific for the malaria protein, Rhop-3, and FL-Rhop-3, were immobilized on the surface of a gold electrode modified with cysteamine. Colloidal gold was used to enhance the detection signal for Rhop-3 antigens. The Rhop-3 antibody was also immobilized on gold electrodes preactivated with dithiobis(succinimidyl proprionate) (DSP). Immobilization was performed at room temperature and at 37 °C. Cyclic voltammetry (CV) was used to monitor the interaction between the immobilized antibody and its cognate antigen in solution, using ferricyanide, K3Fe(CN)6, as reporting electroactive probe. Tests indicate recognition of Rhop-3 protein by the immobilized antibody. Antigen recognition was enhanced by incubation at 37 °C compared with room-temperature incubation. Our results suggest that an immunosensor can be developed and optimized to aid detection of Rhop-3 antigens in samples from malaria patients. As far as we are aware, this is the first amperometric immunosensor targeting Rhop-3 antigen as a malaria biomarker

Predicting Interactive Behavior of Cytokines and Their Receptors by Dielectric Thermal Analysis and Thermogravimetry

Cytokines and soluble cytokine receptors serve as important protein biomarkers for chronic and infectious disease diagnosis. The development of biosensors capable of detecting cytokines or their soluble receptors in patient bodily fluids is a growing area of research. In an ongoing series of studies to understand the thermal analytical behavior of cytokines and their soluble receptors, dielectric thermal analysis (DETA) and thermogravimetry (TG) were used in investigations to determine if differentiations based on dielectric properties (e.g., conductivity) of the proteins could be identified. Permittivity (ε′) and dielectric loss factor (ε′) measurements were performed over a frequency range of 0.1-300,000 Hz. Up to 20 min, water associated with the samples was conductive, interacting with the proteins and affecting the temperature-dependent relaxation spectra of proteins. A trend analysis revealed differences between surface charge at 0.1 Hz and bulk charge at 300,000 Hz. In addition, the greatest change detected among proteins was due to the conductivity (dielectric loss factor). Beyond a 20 min drying time, the observed conductivity was due to intrinsic properties of the proteins with limited dependence on frequency. A 100% water loss was obtained for samples within 20-30 min by TG. Sample drying by TG could serve as a preparatory step in drying protein samples for further DETA and DSC analysis. © 2011 Akadémiai Kiadó, Budapest, Hungary

Human Cytokines Characterized by Dielectric Thermal Analysis, Thermogravimetry, and Differential Scanning Calorimetry

Malaria affects over 500 million people worldwide leading to 1-2 million deaths each year, the majority of whom are children. Four Plasmodium species cause malaria in humans. To properly diagnose, and correctly treat malarial infections, accurate diagnosis of infection is required. Proper diagnosis of infection will result in a reduction of morbidity, mortality, and of drug resistant parasites. However, the current tests for malaria diagnosis do not efficiently identify the appropriate human and parasite biomarkers associated with disease. Detection of specific inflammatory mediators such as cytokines associated with malaria pathogenesis will aid the determination of disease progression, disease prognosis, and the early diagnosis of malaria infection. In this study, we used dielectric thermal analysis (DETA), thermogravimetric analysis, and differential scanning calorimetry (DSC) to characterize five human cytokines (IL-1α, IL-2, IL-4, IL-6, and IL-10), to demonstrate how their thermoanalytical properties can be investigated for sensor design. Analysis for DETA was performed at a frequency range of 0.1-300,000 Hz. Permittivity and loss factor measurements were used to calculate tan δ values. Peak frequencies were used to determine dielectric signatures for each cytokine. The peak frequencies were different for each cytokine analyzed. In addition, activation energies were frequency dependent for IL-2 but frequency independent for the remaining four cytokines. Cytokines were also examined using DSC which established variance in heat of crystallization and heat of fusion of solvent among the five cytokines. A noticeable differentiation was observed with IL-1α among the other four cytokines when analyzed using trend analysis. Detection of unique dielectric signals will aid development of sensitive dielectric sensors capable of detecting cytokines in various human samples. © 2012 Akadémiai Kiadó, Budapest, Hungary

Thermal Behavior and Signature Patterns of Human Cytokine and Soluble Cytokine Receptors Investigated Using Dielectric Thermal Analysis and Thermogravimetry

Cytokines are small regulatory proteins secreted mostly by cells of the immune system. Cytokines participate in anti-inflammatory and pro-inflammatory processes in the body and in responses to host exposure to pathogens. In this study, the thermal behavior of human recombinant cytokines and soluble cytokine receptors; IFNγ, TNFα, IL-1 receptor antagonist, soluble TNF-receptor types 1 and 2, and sIL-2 receptor α were analyzed by dielectric thermal analysis at 37 °C and by thermogravimetry. Measurements were performed at a frequency range of 0.1-300,000 Hz. Permittivity and loss factor measurements were used to calculate mobility of charges (tan δ values) in the proteins from Debye plots. Peak frequencies and polarization times were used to determine dielectric signatures for each cytokine and receptor. Peak frequencies and polarization times were obtained for each cytokine and receptor analyzed. Detection of unique dielectric signatures of the proteins will aid development of sensitive dielectric sensors capable of detecting cytokines and soluble cytokine receptors in various human samples for malaria diagnosis. © 2011 Akadémiai Kiadó, Budapest, Hungary