Immunosensors for quantifying cyclooxygenase 2 pain biomarkers.

BACKGROUND: Cyclooxygenase 2 (COX-2) is a key enzyme in pain biomarkers, inflammation and cancer cell proliferation. Thus biosensors that can quantify pain mediators based on biochemical mechanism are imperative. METHODS: Biomolecular recognition and affinity of antigenic COX-2 with the antibody were investigated using surface plasmon resonance (SPR) and ultra-sensitive portable capillary (UPAC) fluorescence sensors. Polyclonal goat anti-COX-2 (human) antibodies were covalently immobilized on gold SPR surface and direct recognition for the COX-2 antigen assessed. The UPAC sensor utilized an indirect sandwich design involving covalently attached goat anti-COX-2 as the capture antibody and rabbit anti-COX-2 (human) antibody as the secondary antibody. RESULTS: UPAC fluorescence signals were directly proportional to COX-2 at a linear range of 7.46×10⁻4-7.46×10¹ ng/ml with detection limit of 1.02×10⁻4 ng/ml. With SPR a linear range was 3.64×10⁻4-3.64×10² ng/ml was recorded and a detection limit of 1.35×10⁻4 ng/ml. Validation was achieved in simulated blood samples with percent recoveries of 81.39% and 87.23% for SPR and UPAC respectively. CONCLUSION: The developed sensors have the potential to provide objective characterization of pain biomarkers for clinical diagnoses.

Identification and quantitation of Bacillus globigii using metal enhanced electrochemical detection and capillary biosensor.

Presented herein are two detection strategies for the identification and quantification of Bacillus globigii, a spore forming nonpathogenic simulant of Bacillus anthracis. The first strategy involves a label-free, metal-enhanced electrochemical immunosensor for the quantitative detection of Bacillus globigii (atrophaeus). The immunosensor comprises of antibacillus globigii (BG) antibody self-assembled onto a gold quartz crystal electrode via cystamine bond. A solid-phase monolayer of silver underpotentially deposited onto the cystamine modified-Au-electrode surface is used as the redox probe. The monolayer was also generated by adsorbing silver nanoparticles on the gold electrode. When the antibody-modified electrode is exposed to BG spores, the antibody-antigen (Ab-Ag) complex formed insulated the electrode surface toward the silver redox probe. The variation of redox current was found to be proportional to the concentration of the BG spores between 1 x 10(2)-3.5 x 10(4) spores/mL. A detection limit of 602 spores/mL was obtained, which is well-below the infectious dose of anthrax spores at 2.5 x 10(5) spores/mL. The second approach involves the use of ultrasensitive portable capillary biosensor (UPAC) to detect the spores. The capillary is an enclosed system that acts as the flow cell, the waveguide, and the solid support for immobilized bimolecular probes. An evanescent excitation generates a signal from an antigen-antibody-fluorophore complex, which propagates along the capillary and is guided to the detector. A limit of detection of 112 spores/mL was reported using the UPAC sensor. Both methods showed lower detection limits compared to the conventional ELISA. The effect of potential interferants tested using Bacillus pumilus confirmed the selectivity for the analyte. This work should allow the first responders to rapidly detect and quantify Bacillus globigii spores at concentrations that are well-below the infectious dose.