Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Quantum-dot-tagged microbeads for multiplexed optical coding of biomolecules

Abstract

Multicolor optical coding for biological assays has been achieved by embedding different-sized quantum dots (zinc sulfide–capped cadmium selenide nanocrystals) into polymeric microbeads at precisely controlled ratios. Their novel optical properties (e.g., size-tunable emission and simultaneous excitation) render these highly luminescent quantum dots (QDs) ideal fluorophores for wavelength-and-intensity multiplexing. The use of 10 intensity levels and 6 colors could theoretically code one million nucleic acid or protein sequences. Imaging and spectroscopic measurements indicate that the QD-tagged beads are highly uniform and reproducible, yielding bead identification accuracies as high as 99.99% under favorable conditions. DNA hybridization studies demonstrate that the coding and target signals can be simultaneously read at the single-bead level. This spectral coding technology is expected to open new opportunities in gene expression studies, high-throughput screening, and medical diagnostics.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Figure 1: (A) Schematic illustration of optical coding based on wavelength and intensity multiplexing.
Figure 2: Fluorescence micrograph of a mixture of CdSe/ZnS QD-tagged beads emitting single-color signals at 484, 508, 547, 575, and 611 nm.
Figure 3: Quantitative analysis of single-bead signal intensities, uniformity and reproducibility of QD incorporation.
Figure 4: Multicolor QD-tagged beads with precisely controlled fluorescence intensities.
Figure 5: Schematic illustration of DNA hybridization assays using QD-tagged beads.
Figure 6: DNA hybridization assays using multicolor encoded beads.

References

  1. 1

    Fodor, S.P.A. et al. Light-directed, spatially addressable parallel chemical synthesis. Science 251, 767–773 (1991).

    CAS  Article  Google Scholar 

  2. 2

    Schena, M., Shalon, D., Davis, R.W. & Brown, P.O. Quantitative monitoring of gene-expression patterns with a complementary-DNA microarray. Science 270, 467–470 (1995).

    CAS  Article  Google Scholar 

  3. 3

    Dickinson, T.A., Michael, K.L., Kauer, J.S. & Walt, D.R. Convergent, self-encoded bead sensor arrays in the design of an artificial nose. Anal. Chem. 71, 2192–2198 (1999).

    CAS  Article  Google Scholar 

  4. 4

    Clark, H.A., Hoyer, M., Philbert, M.A. & Kopelman, R. Optical nanosensors for chemical analysis inside single living cells: fabrication, characterization, and methods for intracellular delivery of PEBBLE sensors. Anal. Chem. 71, 4831–4836 (1999).

    CAS  Article  Google Scholar 

  5. 5

    Harrison, D.J. et al. Micromachining a miniaturized capillary electrophoresis-based chemical analysis system on a chip. Science 261, 895–897 (1993).

    CAS  Article  Google Scholar 

  6. 6

    Ramsey, J.M., Jacobson, S.C. & Knapp, M.R. Microfabricated chemical measurement systems. Nat. Med. 1, 1093–1096 (1995).

    CAS  Article  Google Scholar 

  7. 7

    Woolley, A.T. & Mathies, R.A. Ultra-high-speed DNA fragment separations using microfabricated capillary array electrophoresis chips. Proc. Natl. Acad. Sci. USA 91, 11348–11352 (1994).

    CAS  Article  Google Scholar 

  8. 8

    Alivisatos, A.P. Semiconductor clusters, nanocrystals, and quantum dots. Science 271, 933–937 (1996).

    CAS  Article  Google Scholar 

  9. 9

    Nirmal, M. & Brus, L.E. Luminescence photophysics in semiconductor nanocrystals. Acc. Chem. Res. 32, 407–414 (1999).

    CAS  Article  Google Scholar 

  10. 10

    Chen, J.W. et al. A microsphere-based assay for multiplexed single nucleotide polymorphism analysis using single base chain extension. Genome Res. 10, 549–557 (2000).

    CAS  Article  Google Scholar 

  11. 11

    Bruchez, M. Jr., Moronne, M., Gin, P., Weiss, S. & Alivisatos, A.P. Semiconductor nanocrystals as fluorescent biological labels. Science 281, 2013–2015 (1998).

    CAS  Article  Google Scholar 

  12. 12

    Chan, W.C.-W. & Nie, S.M. Quantum dot bioconjugates for ultrasensitive nonisotopic detection. Science 281, 2016–2018 (1998).

    CAS  Article  Google Scholar 

  13. 13

    Mitchell, G.P., Mirkin, C.A. & Letsinger R.L. Programmed assembly of DNA functionalized quantum dots. J. Am. Chem. Soc. 121, 8122–8123 (1999).

    CAS  Article  Google Scholar 

  14. 14

    Mattoussi, H. et al. Self-assembly of CdSe-ZnS quantum dots bioconjugates using an engineered recombinant protein. J. Am. Chem. Soc. 122, 12142–12150 (2000).

    CAS  Article  Google Scholar 

  15. 15

    Pathak, S., Choi, S.-K., Arnheim, N. & Thompson, M.E. Hydroxylated quantum dots as luminescent probes for in situ hybridization. J. Am. Chem. Soc. 123, 4103–4104 (2001).

    CAS  Article  Google Scholar 

  16. 16

    Fulton, R.J., McDade, R.L., Smith, P.L., Kienker, L.J. & Kettman, J.R. Jr. Advanced multiplexed analysis with the FlowMetrix (TM) system. Clin. Chem. 43, 1749–1756 (1997).

    CAS  PubMed  PubMed Central  Google Scholar 

  17. 17

    Steemers, F.J., Ferguson, J.A. & Walt, D.R. Screening unlabeled DNA targets with randomly ordered fiber-optic gene arrays. Nat. Biotechnol. 18, 91–94 (2000).

    CAS  Article  Google Scholar 

  18. 18

    Ferguson, J.A., Boles, T.C., Adams, C.P. & Walt, D.R. A fiber-optic DNA biosensor microarray for the analysis of gene expression. Nat.re Biotechnol. 14, 1681–1684 (1996).

    CAS  Article  Google Scholar 

  19. 19

    Ferguson, J.A., Steemers, F.J. & Walt, D.R. High-density fiber-optic DNA random microsphere arrays. Anal. Chem. 72, 5618–5624 (2000).

    CAS  Article  Google Scholar 

  20. 20

    Mandecki, W., Ernst, E. & Kogan, N. Light-powered microtransponders for high multiplex-level analyses of nucleic acids. Abstr. Pap. Am. Chem. Soc. 219, U755–U755, Part 1 (2000).

    Google Scholar 

  21. 21

    Moran, E.J. et al. Radio-frequency tag encoded combinatorial library method for the discovery of tripeptide-substituted cinnamic acid inhibitors of the protein-tyrosine-phosphatase PTP1B. J. Am. Chem. Soc. 117, 10787–10788 (1995).

    CAS  Article  Google Scholar 

  22. 22

    Hines, M.A. & Guyot-Sionnest, P. Synthesis and characterization of strongly luminescing ZnS-capped CdSe nanocrystals. J. Phys. Chem. 100, 468–471 (1996).

    CAS  Article  Google Scholar 

  23. 23

    Dabbousi, B.O. et al. (CdSe)ZnS core-shell quantum dots: synthesis and characterization of a size series of highly luminescent nanocrystallites. J. Phys. Chem. B 101, 9463–9475 (1997).

    CAS  Article  Google Scholar 

  24. 24

    Peng, X., Schlamp, M.C., Kadavanich, A.V. & Alivisatos, A.P. Epitaxial growth of highly luminescent CdSe/CdS core/shell nanocrystals with photostability and electronic accessibility. J. Am. Chem. Soc. 119, 7019–7029 (1997).

    CAS  Article  Google Scholar 

  25. 25

    Zhang, Y.-Z., Kemper, C.R. & Haugland, R.P. Microspheres with fluorescent spherical zones. US 5,786,219 (1998).

    Google Scholar 

  26. 26

    Kagan, C.R., Murray, C.B., Nirmal, M. & Bawendi, M.G. Electronic energy transfer in CdSe quantum dot solids. Phys. Rev. Lett. 76, 1517–1520 (1996).

    CAS  Article  Google Scholar 

  27. 27

    Micic, O.I., Jones, K.M., Cahill, A. & Nozik, A.J. Optical, electronic, and structural properties of uncoupled and close-packed arrays of InP quantum dots. J. Phys. Chem. B 102, 9791–9796 (1998).

    CAS  Article  Google Scholar 

  28. 28

    Prummer, M. et al. Single-molecule identification by spectrally and time-resolved fluorescence detection. Anal. Chem. 72, 443–447 (2000).

    CAS  Article  Google Scholar 

  29. 29

    Dorsey, J.G. & Cooper, W.T. Retention mechanisms of bonded-phase liquid-chromatography. Anal. Chem. 66, 857A–867A (1994).

    CAS  Article  Google Scholar 

  30. 30

    Fu, A.Y., Spence, C., Scherer. A., Arnold, F.H & Quake, S.R. A microfabricated fluorescence-activated cell sorter. Nat. Biotechnol. 17, 1109–1111 (1999).

    CAS  Article  Google Scholar 

  31. 31

    Miraglia. S. et al. Homogeneous cell- and bead-based assays for high throughput screening using fluorometric microvolume assay technology. J. Biomol. Screening 4, 193–204 (1999).

    CAS  Article  Google Scholar 

  32. 32

    Hermkens, P.H.H., Ottenheijm, H.C.J. & Rees, D. Solid-phase organic reactions: a review of the recent literature. Tetrahedron 52, 4527–4554 (1996).

    CAS  Article  Google Scholar 

  33. 33

    Yang, B.Z., Chen, L.W. & Chiu, W.Y. Effects of acrylic acid on number density of polymer particles in emulsifier-free emulsion copolymerization of styrene and acrylic acid. Polymer J. 29, 737–743 (1997).

    CAS  Article  Google Scholar 

  34. 34

    Hermanson, G.T. Bioconjugate techniques. (Academic Press, New York; 1996).

    Google Scholar 

Download references

Acknowledgements

We are grateful to Warren C.-W. Chan for help in quantum dot synthesis and for stimulating discussions. This work was supported in part by the National Institutes of Health and the Department of Energy. S.N. acknowledges the Whitaker Foundation for a Biomedical Engineering Award and the Beckman Foundation for a Beckman Young Investigator Award.

Author information

Affiliations

Authors

Corresponding author

Correspondence to Shuming Nie.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Han, M., Gao, X., Su, J. et al. Quantum-dot-tagged microbeads for multiplexed optical coding of biomolecules. Nat Biotechnol 19, 631–635 (2001). https://doi.org/10.1038/90228

Download citation

Further reading

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing