Simultaneous multicolor detection system of the single-molecular microbial antigen by total internal reflection fluorescence microscopy with fluorescent nanocrystal quantum dots

Akiyoshi Hoshino; Kouki Fujioka; Mayu Yamamoto; Noriyoshi Manabe; Masato Yasuhara; Kazuo Suzuki; Kenji Yamamoto M.D.

doi:10.1117/12.630344

21 November 2005 Simultaneous multicolor detection system of the single-molecular microbial antigen by total internal reflection fluorescence microscopy with fluorescent nanocrystal quantum dots

Akiyoshi Hoshino, Kouki Fujioka, Mayu Yamamoto, Noriyoshi Manabe, Masato Yasuhara, Kazuo Suzuki, Kenji Yamamoto M.D.

Author Affiliations +

Proceedings Volume 6009, Optical Methods in Drug Discovery and Development; 60090P (2005) https://doi.org/10.1117/12.630344
Event: Optics East 2005, 2005, Boston, MA, United States

Abstract

Immunological diagnostic methods have been widely performed and showed high performance in molecular and cellular biology, molecular imaging, and medical diagnostics. We have developed novel methods for the fluorescent labeling of several antibodies coupled with fluorescent nanocrystals QDs. In this study we demonstrated that two bacterial toxins, diphtheria toxin and tetanus toxin, were detected simultaneously in the same view field of a cover slip by using directly QD-conjugated antibodies. We have succeeded in detecting bacterial toxins by counting luminescent spots on the evanescent field with using primary antibody conjugated to QDs. In addition, each bacterial toxin in the mixture can be separately detected by single excitation laser with emission band pass filters, and simultaneously in situ pathogen quantification was performed by calculating the luminescent density on the surface of the cover slip. Our results demonstrate that total internal reflection fluorescence microscopy (TIRFM) enables us to distinguish each antigen from mixed samples and can simultaneously quantitate multiple antigens by QD-conjugated antibodies. Bioconjugated QDs could have great potentialities for in practical biomedical applications to develop various high-sensitivity detection systems.

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Akiyoshi Hoshino, Kouki Fujioka, Mayu Yamamoto, Noriyoshi Manabe, Masato Yasuhara, Kazuo Suzuki, and Kenji Yamamoto M.D. "Simultaneous multicolor detection system of the single-molecular microbial antigen by total internal reflection fluorescence microscopy with fluorescent nanocrystal quantum dots", Proc. SPIE 6009, Optical Methods in Drug Discovery and Development, 60090P (21 November 2005); https://doi.org/10.1117/12.630344

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