Carp vitellogenin detection by an optical waveguide lightmode spectroscopy biosensor.

A label-free carp vitellogenin sensor has a strong potential for on-site monitoring on the possible contamination of edible fish with endocrine disruptors as a sum parameter in an inland carp farm. In this study, we performed a sensitive detection for carp vitellogenin with a direct-binding optical waveguide lightmode spectroscopy-based immunosensor. Carp vitellogenin bound over the sensor surface quite specifically, judging from the sensor responses according to stepwise antibody immobilization. This was also supported by a negligible sensor response found at bovine serum albumin immobilization. When plotted in double-logarithmic scale for carp vitellogenin concentrations of 0.00675-67.5 nM, a linear relationship was found between analyte concentration and sensor response, together with the limit of detection of 0.00675 nM. The reusability of the immunosensor after the regeneration with 10mM HCl was reasonably good, as presumed from the coefficient of variability of 6.02% for nine repetitive measurements. The model sample prepared by spiking a purified carp vitellogenin into a 10-fold diluted vitellogenin-free carp serum in 9.45 nM showed the response ratio of 96.70% against 9.45 nM of the purified carp vitellogenin. When a female and male carp sera induced with 17beta-estradiol injection were analyzed, biomarker induction was even identifiable at 2000-fold serum dilution.

Sulfamethazine detection with direct-binding optical waveguide lightmode spectroscopy-based immunosensor.

A direct-binding optical waveguide lightmode spectroscopy-based immunosensor detecting sulfamethazine (SMZ) was prepared, followed by the measurement of its specificity and sensitivity. System construction was undertaken with a peristaltic pump, an injector and the main unit comprising a sensor holder, two signal-harvesting photodiodes, a beam mirror, shutter and He-Ne laser source emitting a monochrome light (λ=632.8nm), plus a PC. Antibody immobilization was performed in situ by covalent binding of an anti-SMZ antibody over the surface of a glutaraldehyde-activated 3-aminopropyltriethoxysilane-treated sensor chip. The reaction buffer for the system was 4mM Tris-HCl (pH 7.2) that showed a medium surface coverage and stable baseline. Sensor response was quite specific to antibody-antigen complexation, as judged from no sensor response caused by bovine serum albumin immobilization. The sensor responses according to SMZ concentrations from 10(-8) to 10(-2)M increased linearly in a semi-logarithmic scale, with the limit of detection of 10(-8)M. The immunosensor was favorably reusable for SMZ screening.

High-sensitivity detection for model organophosphorus and carbamate pesticide with quartz crystal microbalance-precipitation sensor.

The effect of acetylcholinesterase (AChE) immobilization over the surface of a quartz crystal microbalance (QCM), by chemisorption of the AChE thiolated with a heterobifunctional cross-linker, sulfo-succinimidyl-6-[3-(2-pyridyldithio)propionamido]hexanoate, and carboxyl-amine coupling of AChE to 3-mercaptopropionic acid self-assembled monolayer, on the responses of a batch-type QCM-precipitation sensor was compared, resulting in a better sensitivity and binding efficiency in the former method. When an inhibition study with the developed sensor was undertaken at the optimized AChE immobilization with varying concentrations of a model organophosphorus pesticide EPN and carbamate one carbofuran, a sensitive detection for them was possible with the limit of detection corresponding to 1.55 x 10(-8) and 1.30 x 10(-9)M, respectively.

Development of a direct-binding chloramphenicol sensor based on thiol or sulfide mediated self-assembled antibody monolayers.

A batch-type antibody-immobilized quartz crystal microbalance (QCM) system for detecting chloramphenicol (CAP) was developed. To bind an anti-CAP antibody onto the gold electrode surface of piezoelectric crystals, self-assembled monolayers (SAMs) of different thiols or sulfides were formed by a chemisorption procedure. Then, the anti-CAP antibody was covalently linked to the pre-formed monolayers by an activation procedure using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride and N-hydroxysulfosuccinimide. The antibody-immobilized QCM chip thus prepared was installed in a well holder and was measured for sensor response. Compared with the bare QCM chip and the QCM chip only coated with 3-mercaptopropionic acid (MPA), the antibody-immobilized sensor showed greatly enhanced frequency shifts by 10-50-fold after CAP injection. In this case, CAP detection which was indicated by steady-state resonant frequency shift was accomplished within 10 min. When CAP solution was injected into the reaction cell in 50mM concentration, the frequency shifts obtained were, respectively, 530 and 505 Hz in case of thiosalicylic acid and MPA immobilization. Repeated use of the sensor chips up to eight times was possible after 1 min regeneration with 0.1M NaOH. This system demonstrated a potential application of thiol or sulfide mediated SAMs as the pre-coatings of a real-time detection on CAP in solution.