DEVELOPMENT OF LABEL-FREE BIOSENSOR FOR DETECTING STEROID HORMONE CONCENTRATION IN FISH

A novel label-free immunosensor for detecting steroid hormone was developed. The principle of the sensor system is based on differences in the electrochemical activity induced by an immunoreaction that depends on the levels of steroid hormone in the sample. A gold electrode functionalized with 3-mercaptopropionic acid (MPA) self- assembled monolayer was used to fabricate electrochemical immunosensor. In addition, single-wall carbon nanotube (SWCNT) was selected to expandthe dynamic range of the sensor. The sensor was immersed into a sample solution and measurements were determined by cyclic voltammetry. Each electrochemical measurement including sample immerse only took about 15min. In this study, we introduced an application of our sensor in detecting 17, 20β- dihydroxy-4-pregnen- 3-one (DHP). The immunosensor showed a specific response to DHP, and the oxidation peak current linearly decreased in the range of 7.8-500.0pg ml-1 (without SWCNT) and 15.6-50000.0pg ml-1 (with SWCNT). The sensor system was then applied to monitor DHP of goldfish (Carassiusauratus) and was compared with the levels of the same samples determined using ELISA as the convention method. Blood plasma of fish was collected every 3h after administering a DHP inducer. A good relationship (coefficient: 0.934) was observed between DHP levels determined by both methods. Keyword: Biosensor, Steroid hormone, Immunoassay, Fish, Single-walled carbon nanotube

Wireless Biosensor System for Real-Time L-Lactic Acid Monitoring in Fish

We have developed a wireless biosensor system to continuously monitor L-lactic acid concentrations in fish. The blood L-lactic acid level of fish is a barometer of stress. The biosensor comprised Pt-Ir wire (φ0.178 mm) as the working electrode and Ag/AgCl paste as the reference electrode. Lactate oxidase was immobilized on the working electrode using glutaraldehyde. The sensor calibration was linear and good correlated with L-lactic acid levels (R = 0.9959) in the range of 0.04 to 6.0 mg·dL<sup>−1</sup>. We used the eyeball interstitial sclera fluid (EISF) as the site of sensor implantation. The blood L-lactic acid levels correlated closely with the EISF L-lactic acid levels in the range of 3 to 13 mg·dL<sup>−1</sup> (R = 0.8173, n = 26). Wireless monitoring of L-lactic acid was performed using the sensor system in free-swimming fish in an aquarium. The sensor response was stable for over 60 h. Thus, our biosensor provided a rapid and convenient method for real-time monitoring of L-lactic acid levels in fish

Ultra highly sensitive method for detecting Edwardsiella ictaluri using high-gradient immunomagnetic separation with polymerase chain reaction

Edwardsiella ictaluri causes an economically important bacterial disease in farm-raised catfish in the USA and abroad. To elucidate the route of infection for fish bacterial disease, it is important to monitor both fish health and trace bacteria in the water environment. In this study, we applied an ultra highly sensitive method to detect E. ictaluri using high-gradient immunomagnetic separation (HGIMS) with polymerase chain reaction (PCR). HGIMS is a magnetic separation method in which the magnetic force is strengthened by integrating a magnetic gradient between the magnetic filter and nearby column. Immunomagnetic beads were specifically designed to react with the target bacteria, allowing for more efficient collection. The accumulated beads were released from the filter by releasing the magnetic force. After the process, DNA was extracted from the concentrated cells, and PCR was applied to detect E. ictaluri. The HGIMS system had higher detection sensitivity using than the conventional method, and the total assay time, including sample preparation, was about 3.5h. The optimal reaction time of immunomagnetic beads was 15min and the flow rate of the HGIMS system was 10mlmin−1. PCR products of the expected size were obtained from samples concentrated up to 102cfuml−1. The proposed system appears to be suitable for highly sensitive detection of E. ictaluri. Improvements in the bacteria recovery ability of the immunomagnetic beads will further increase the detection limits. Keywords: Edwardsiella ictaluri, Magnetic separation, Polymerase chain reaction, Fish disease, Bacteria detection, Bioanalytical metho

Development Of Label-free Biosensor For Detecting Steroid Hormone Concentration In Fish

A novel label-free immunosensor for detecting steroid hormone was developed. The principle of the sensor system is based on differences in the electrochemical activity induced by an immunoreaction that depends on the levels of steroid hormone in the sample. A gold electrode functionalized with 3-mercaptopropionic acid (MPA) self- assembled monolayer was used to fabricate electrochemical immunosensor. In addition, single-wall carbon nanotube (SWCNT) was selected to expandthe dynamic range of the sensor. The sensor was immersed into a sample solution and measurements were determined by cyclic voltammetry. Each electrochemical measurement including sample immerse only took about 15min. In this study, we introduced an application of our sensor in detecting 17, 20β- dihydroxy-4-pregnen- 3-one (DHP). The immunosensor showed a specific response to DHP, and the oxidation peak current linearly decreased in the range of 7.8-500.0pg ml-1 (without SWCNT) and 15.6-50000.0pg ml-1 (with SWCNT). The sensor system was then applied to monitor DHP of goldfish (Carassiusauratus) and was compared with the levels of the same samples determined using ELISA as the convention method. Blood plasma of fish was collected every 3h after administering a DHP inducer. A good relationship (coefficient: 0.934) was observed between DHP levels determined by both methods