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A new microvolume technique for bioaffinity assays using two-photon excitation

Nature Biotechnology volume 18, pages 548–550 (2000)Cite this article

Abstract

Bioaffinity binding assays such as the immunoassay are widely used in life science research. In an immunoassay, specific antibodies are used to bind target molecules in the sample, and quantification of the binding reaction reveals the amount of the target molecules. Here we present a method to measure bioaffinity assays using the two-photon excitation of fluorescence. In this method, microparticles are used as solid phase in binding the target molecules. The degree of binding is then quantified from individual microparticles by use of two photon excitation of fluorescence. We demonstrated the effectiveness of the method using the human α-fetoprotein (AFP) immunoassay, which is used to detect fetal disorders. The sensitivity and dynamic range we obtained with this assay indicate that this method can provide a cost-effective and simple way to measure various biomolecules in solution for research and clinical applications.

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Figure 1: Optical set-up of the instrument.
Figure 2: A single-particle signal trace at 400 ng ml−1 of AFP from the confocal scattering channel (black) and fluorescence channel (gray).
Figure 3: Intensity distribution histograms of three different particle categories.
Figure 4: AFP background created response curves for a separation-free single-step assay measured in two volumes: 25 μl (▴) and 2 μl (○).

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Acknowledgements

This work was supported by the Academy of Finland and National Technology Development Agency, TEKES. Special thanks to Mr. Honda of Fujirebio Inc. (Tokyo, Japan) and Mr. Nasu and Mr. Sakurai of SRL Inc. (Tokyo, Japan) for supplying, preparing, and characterizing the material for AFP assay.

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  1. Laboratory of Biophysics, Institute of Biomedicine, University of Turku, P.O. Box 123, 20521, Turku, Finland

    Pekka Hänninen, Aleksi Soini, Niko Meltola, Juhani Soini, Jori Soukka & Erkki Soini

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  1. Pekka Hänninen
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Correspondence to Pekka Hänninen.

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Hänninen, P., Soini, A., Meltola, N. et al. A new microvolume technique for bioaffinity assays using two-photon excitation. Nat Biotechnol 18, 548–550 (2000). https://doi.org/10.1038/75421

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