Biocapteurs et bioélectronique

A novel detection system for biological agents using Loop-Mediated Isothermal Amplification (LAMP) and an electrochemical DNA chip, Bio BulwarkTM, has been developed. Genomic DNAs/RNAs roughly extracted from biological agents were amplified by the LAMP method using target-specific primers and strand-displace polymerase. The amplified DNAs were processed to hybridization with the electrochemical DNA chip. Anodic current derived from hoechst 33258, an electrochemically active intercalator, was relative to the amount of the target DNA hybridized with target-specific probes on the DNA chip. All reactions and measurements were conducted using disposable cassettes, which included the reagents for amplification and the DNA chip, and a mobile automatic detection system. The preliminary study showed that accurate detection of B. anthracis was carried out within 70 minutes. This system would prevent or minimize damage in the event of a criminal attack involving a biological threat.

This paper first time describes Surface Plasmon Resonance (SPR) based immuno-sensing system for detection of fungal teliospores of Karnal Bunt (KB) disease of wheat incited by Tilletia indica. The results of present study were compared with the results of lab scale ELISA developed earlier in our lab. The approach involves the use of a rabbit polyclonal (anti-teliospore) antibody and a SPR sensor for label-free detection of teliospores from the model organism Tilletia indica. When the interaction between binder (anti-teliospore antibody) and teliosporic antigen(s) as analyte occurs at the sensor surface, a sensorgram was obtained in real time by plotting the signal against time. Under optimised assay conditions as few as 5 teliospores were detected by ELISA which justifies the greater affinity of raised anti-teliospore antibodies for its antigen. The sensitivity of each immunosensor constructed with
the different amounts of antigen solution is determined by sensogram analysis showed detection sensitivity as low as 625 pg equivalent to 2.5 teliospores over the sensor surface by getting an angle of dip indicating greater affinity of raised anti-teliospore antibodies. Thus, SPR based affinity sensor showed higher sensitivity due to greater signal response. The results of cross reactivity studies using different related fungal spores/teliospores showed almost differential SPR response indicating cross reaction using immunosensing system. The cross-reactivity of anti-teliospore antibodies with related fungal spores/teliospores can be explained due to its polyclonal nature and also due to antigenic homology amongst pathogens used in present study.

β-thalassemias are very heterogeneous in nature, with respect to the molecular basis of the disease. This type of disorder is caused by extensive deletions occurring within the β-globin gene locus, present on chromosome number 11. Due to point mutations, stop codons in the β-globin mRNA is introduced, thus curbing its normal function. This paper reviews conventional and latest β-thalassemia detection techniques. Amongst the conventional techniques, the DNA-based piezoelectric biosensors were based on the observation of distinguishable frequency shifts, resulting from the hybridisation between an oligonucleotide probe which was immobilized on a gold electrode of a quartz crystal and the complementary strand; present in solution. Q-primer real time PCR (Q-PCR) system utilises
the 5’>3’ exonuclease activity of a DNA polymerase, which cleaves the nucleotides and results in fluorescence. Amongst the latest detection methods, the capacitive micro-membrane and surface stress based biosensors require flexible structures, microcantilevers or membranes, where one of their surfaces is functionalized with the probe biomolecules. The interaction with the appropriate target molecules induces surface stress variations and finally changes in the deflection of the structure. In bead based biosensors, the natural immobilization of the beads into the polyacrylamide gel pads allows the beads to acquire unique spatial addresses that is recorded via image, and thus helps in avoiding labeling or color encoding steps. These techniques are more efficient and overcome the disadvantages of conventional techniques that were costly, less sensitive and complex too.

A bioanode for glucose based biofuel cell have been fabricated by combining glucose oxidase and highly ordered Palladium (Pd) nanowire array electrode. The Pd nanowires were 5.57 μm in length and 64.28 nm in diameter. The hydrogel composite modified Pd nanowire array bioanodes were characterized with cyclic voltammetry in the presence of different scan rates and different substrate concentrations under physiological conditions. The electrochemical measurements showed a linear amperometric response in the range of 1 mM to 13 mM. In addition, surface and structural characterization were performed by scanning electron microscopy leading to the conclusion that the Pd nanowires are highly ordered and vertically aligned and thereby promotes the number of electroactive sites for the catalysis of the oxidation of glucose. The combination of glucose oxidase and Pd nanowire arrays revealed the capability of these enzymatic bioanodes to perform direct electron transfer to Pd nanostructure surfaces and be used in a glucose biofuel cell.

Measurements of serum cholesterol levels are important in the diagnosis and classification of hyper
lipoproteinemias. Elevated cholesterol levels may occur with hypothyroidism, nephrotic syndrome, diabetes, and various liver diseases. The present investigation was carried out for the development of rapid, highly sensitive and economic cholesterol biosensor for the determination of blood cholesterol. The PVP encapsulated ZnS nanoparticals have been synthesized and characterized by Fourier Transform InfraRed (FTIR) spectroscopy, UV-visible spectroscopy, and Atomic force microscopy techniques in the present investigation. The size of the nanoparticles is found to be ranging between 21-22 nm.

The surface morphology of the sensing area of functionalized MWCNT was examined using Scanning Electron Microscopy (SEM). The length of the MWCNTs is found to be approximately 2 microns. In the IR spectrum of functionalized MWCNTs, the presence of bands at 1636 cm-1 and 3434 cm-1 confirmed the presence of carbonyl and hydroxyl moieties of carboxylic acid group.

Cholesterol Oxidase was immobilized on to the ZnS nanoparticles and Multiwall Carbon Nanotubes (MWCNTs) which they were placed on a glassy carbon electrode surface using Nafion by LBL assembly technique. Cyclic voltammetric study reveals that the fast electron transfer between electrodes was achieved through the incorporation of ChOx into the Nafion-ZnS-MWCNTs film. ZnS and MWCNTs played an important role in facilitating the electron transfer between the ChOx and the electrode surface.

The cholesterol  bioelectrode shows detection range of 10-450 mg/dl and good linearity is obtained in 50-450 mg/dl (1.3 -11.6 mM) range with linear regression coefficient 'R2' as 0.986. The Michaelis-Menten constant Km value obtained as 0.84 mM using Lineweaver-Burke plot by plotting 1/V vs 1/Conc. The low Km value  0.84 mM indicates high affinity of immobilized ChOx to cholesterol. It exhibits optimum pH 7.0 and optimum temperature 35°C. Therefore, the bioelectrode fabricated in this study is promising for cholesterol detection in human blood.

Bioimpedance analysis could provide a clear figure about changes in cells and tissues based on frequencydependent changes, due to their electrical resistances for the applied electrical current. This review explains the physical principle of bioimpedance. And to monitor the progression of radiation induced tissue injury, particularly in radiotherapies. The interaction of radiation with the biological tissues and a prediction for their earlier and later alterations due to exposure are discussed. As well as, an overview for tissue identification by bioelectrical impedance analysis (BIA) is proposed. Bioimpedance analysis “applications and its limitations” in the health care, clinical practice and prognosis of overall survival in cancer patients are discussed.