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Fiber-Optic Fluoroimmunoassay System with a Flow-Through Cell for Rapid On-Site Determination of Escherichia coli O157:H7 by Monitoring Fluorescence Dynamics.

Miyajima K, Koshida T, Arakawa T, Kudo H, Saito H, Yano K, Mitsubayashi K - Biosensors (Basel) (2013)

Bottom Line: The measurement for each sample was completed within 12 min.This minimized the time for measurement down to 6 min.The system is suitable for rapid and direct determination for microorganisms or bacteria in food, clinical, and environmental sources.

View Article: PubMed Central - PubMed

Affiliation: Department of Advanced Sciences and Technology for Biomedical Sensors, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8549, Japan; E-Mail: miya.bdi@tmd.ac.jp ; Department of Biomedical Devices and Instrumentation, Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University, 2-3-10 Kanda-Surugadai, Chiyoda-ku, Tokyo 101-0062, Japan; E-Mails: arakawa.bdi@tmd.ac.jp (T.A.); kudo.bdi@tmd.ac.jp (H.K.).

ABSTRACT
Dynamic fluoroimmunoassay with a flow-through system using optical fiber probes consisting of polystyrene was developed and applied to a quantitative detection of E. coli O157:H7. The system measures E. coli as fluorescence of sandwich-type immune complexes formed by capture antibodies immobilized on the surface of the probe, E. coli cells, and fluorescently labeled detection antibodies. Excitation was carried out using an evanescent wave from the probe. Resulting fluorescence recoupled into the probe was detected by a photodiode. The assay system was constructed with a flow cell which was available for sequential injection of experimental reagents. In vitro characterization was performed using the flow cell, and the calibration range of E. coli O157:H7 was from 10(3) to 10(7) cells/mL. The measurement for each sample was completed within 12 min. Furthermore, it was also possible to estimate the concentrations of E. coli O157:H7 by the increasing rate of fluorescence during binding reaction of detection antibodies to antigens. This minimized the time for measurement down to 6 min. The system is suitable for rapid and direct determination for microorganisms or bacteria in food, clinical, and environmental sources.

No MeSH data available.


Changes in fluorescent intensities of response to E. coli O157:H7 concentrations. The signal of nonspecific reaction was negligibly low at step II, and stable current values at every four measurement steps were obtained immediately.
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biosensors-03-00120-f004: Changes in fluorescent intensities of response to E. coli O157:H7 concentrations. The signal of nonspecific reaction was negligibly low at step II, and stable current values at every four measurement steps were obtained immediately.

Mentions: Figure 4 shows the variation of fluorescent intensity corresponding to E. coli O157:H7 concentrations at each measurement step. No significant nonspecific reaction was confirmed at step II, and the current was sufficiently stable throughout the measurement. As the E. coli O157:H7 concentration increased, the fluorescence signal also increased. Calibration curve measured by the flow-through measurement system is shown in Figure 5. The calibration curve was made from data which had fluorescent difference between steps III and IV. As a result, the highest fluorescence was obtained at 107 cells/mL, and the lower detection limit was determined to be 103 cells/mL. The coefficient of determination (R2) was 0.999 deduced by regression analysis as shown by the following equation:(1)


Fiber-Optic Fluoroimmunoassay System with a Flow-Through Cell for Rapid On-Site Determination of Escherichia coli O157:H7 by Monitoring Fluorescence Dynamics.

Miyajima K, Koshida T, Arakawa T, Kudo H, Saito H, Yano K, Mitsubayashi K - Biosensors (Basel) (2013)

Changes in fluorescent intensities of response to E. coli O157:H7 concentrations. The signal of nonspecific reaction was negligibly low at step II, and stable current values at every four measurement steps were obtained immediately.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4263595&req=5

biosensors-03-00120-f004: Changes in fluorescent intensities of response to E. coli O157:H7 concentrations. The signal of nonspecific reaction was negligibly low at step II, and stable current values at every four measurement steps were obtained immediately.
Mentions: Figure 4 shows the variation of fluorescent intensity corresponding to E. coli O157:H7 concentrations at each measurement step. No significant nonspecific reaction was confirmed at step II, and the current was sufficiently stable throughout the measurement. As the E. coli O157:H7 concentration increased, the fluorescence signal also increased. Calibration curve measured by the flow-through measurement system is shown in Figure 5. The calibration curve was made from data which had fluorescent difference between steps III and IV. As a result, the highest fluorescence was obtained at 107 cells/mL, and the lower detection limit was determined to be 103 cells/mL. The coefficient of determination (R2) was 0.999 deduced by regression analysis as shown by the following equation:(1)

Bottom Line: The measurement for each sample was completed within 12 min.This minimized the time for measurement down to 6 min.The system is suitable for rapid and direct determination for microorganisms or bacteria in food, clinical, and environmental sources.

View Article: PubMed Central - PubMed

Affiliation: Department of Advanced Sciences and Technology for Biomedical Sensors, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8549, Japan; E-Mail: miya.bdi@tmd.ac.jp ; Department of Biomedical Devices and Instrumentation, Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University, 2-3-10 Kanda-Surugadai, Chiyoda-ku, Tokyo 101-0062, Japan; E-Mails: arakawa.bdi@tmd.ac.jp (T.A.); kudo.bdi@tmd.ac.jp (H.K.).

ABSTRACT
Dynamic fluoroimmunoassay with a flow-through system using optical fiber probes consisting of polystyrene was developed and applied to a quantitative detection of E. coli O157:H7. The system measures E. coli as fluorescence of sandwich-type immune complexes formed by capture antibodies immobilized on the surface of the probe, E. coli cells, and fluorescently labeled detection antibodies. Excitation was carried out using an evanescent wave from the probe. Resulting fluorescence recoupled into the probe was detected by a photodiode. The assay system was constructed with a flow cell which was available for sequential injection of experimental reagents. In vitro characterization was performed using the flow cell, and the calibration range of E. coli O157:H7 was from 10(3) to 10(7) cells/mL. The measurement for each sample was completed within 12 min. Furthermore, it was also possible to estimate the concentrations of E. coli O157:H7 by the increasing rate of fluorescence during binding reaction of detection antibodies to antigens. This minimized the time for measurement down to 6 min. The system is suitable for rapid and direct determination for microorganisms or bacteria in food, clinical, and environmental sources.

No MeSH data available.