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An ultrasensitive universal detector based on neutralizer displacement

Abstract

Diagnostic technologies that can provide the simultaneous detection of nucleic acids for gene expression, proteins for host response and small molecules for profiling the human metabolome will have a significant advantage in providing comprehensive patient monitoring. Molecular sensors that report changes in the electrostatics of a sensor's surface on analyte binding have shown unprecedented sensitivity in the detection of charged biomolecules, but do not lend themselves to the detection of small molecules, which do not carry significant charge. Here, we introduce the neutralizer displacement assay that allows charge-based sensing to be applied to any class of molecule irrespective of the analyte charge. The neutralizer displacement assay starts with an aptamer probe bound to a neutralizer. When analyte binding occurs the neutralizer is displaced, which results in a dramatic change in the surface charge for all types of analytes. We have tested the sensitivity, speed and specificity of this system in the detection of a panel of molecules: (deoxy)ribonucleic acid, ribonucleic acid, cocaine, adenosine triphosphate and thrombin.

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Figure 1: Summary of the NDA and sensor chips utilized for testing.
Figure 2: NDA small molecule detection: ATP and cocaine.
Figure 3: NDA nucleic acid and bacterial detection.
Figure 4: NDA protein detection.

References

  1. Xia, F. et al. Colorimetric detection of DNA, small molecules, proteins, and ions using unmodified gold nanoparticles and conjugated polyelectrolytes. Proc. Natl Acad. Sci. USA 107, 10837–10841 (2010).

    Article  CAS  Google Scholar 

  2. Zhang, M., Win, B. C., Tan, W. & We, B. C. A versatile graphene-based fluorescence ‘on/off’ switch for multiplex detection of various targets. Biosens. Bioelectron. 26, 3260–3265 (2011).

    Article  CAS  Google Scholar 

  3. Drummond, T. G., Hill, M. G. & Barton, J. K. Electrochemical DNA sensors. Nature Biotechnol. 21, 1192–1199 (2008).

    Article  Google Scholar 

  4. Clack, N. G., Asalaita, K. & Groves, J. T. Electrostatic readout of DNA microarrays with charged microspheres. Nature Biotechnol. 26, 825–830 (2008).

    Article  CAS  Google Scholar 

  5. Morrow, T. J., Li, M., Kim, J., Mayer, T. S. & Keating, C. D. Programmed assembly of DNA-coated nanowire devices. Science 323, 352 (2009).

    Article  CAS  Google Scholar 

  6. Zheng, G., Patolsky, F., Cui, Y., Wang, U. W. & Lieber, C. M. Multiplexed electrical detection of cancer markers with nanowire sensors arrays. Nature Biotechnol. 23, 1294–1300 (2005).

    Article  CAS  Google Scholar 

  7. Patolsky, F., Zheng, G. & Lieber, C. M. Fabrication of silicon nanowire devices for ultrasensitive, label-free, real-time detection of biological and chemical species. Nature Protocols 1, 1711–1724 (2006).

    Article  CAS  Google Scholar 

  8. Tian, B. et al. Three-dimensional, flexible nanoscale field-effect transistors as localized bioprobes. Science 13, 830–834 (2010).

    Article  Google Scholar 

  9. Zheng. G., Gao, X. P. A. & Lieber, C. M. Frequency domain detection of biomolecules using silicon nanowire biosensors. Nano Lett. 10, 3179–3183 (2010).

    Article  CAS  Google Scholar 

  10. Wu, G et al. Bioassay of prostate-specific antigen (PSA) using microcantilevers. Nature Biotechnol. 19, 856–860 (2001).

    Article  CAS  Google Scholar 

  11. Fritz, J. et al. Translating biomolecular recognition into nanomechanics. Science 288, 316–318 (2000).

    Article  CAS  Google Scholar 

  12. Xiang, Y. & Lu, Y. Using personal glucose meters and functional DNA sensors to quantify a variety of analytical targets. Nature Chem. 3, 697–670 (2011).

    Article  CAS  Google Scholar 

  13. Cheng, A. K. H., Ge, B. X. & Yu, H. Z. Label-free voltammetric detection of lysozyme with aptamer-modified gold electrodes. Anal. Chem. 79, 5158–5164 (2007).

    Article  CAS  Google Scholar 

  14. Baker, B. R. et al. An electronic, aptamer-based small molecule sensor for the rapid, reagentless detection of cocaine in adulterated samples and biological fluids. J. Am. Chem. Soc. 128, 3138–3139 (2006).

    Article  CAS  Google Scholar 

  15. Xiao, Y., Lai, R. Y. & Plaxco, K. W. Preparation of electrode-immobilized, redox-modified oligonucleotides for electrochemical DNA and aptamer-based sensing. Nature Protocols 2, 2875–2880 (2007).

    Article  CAS  Google Scholar 

  16. Cash, K. J., Ricci, F. & Plaxco, K. W. An electrochemical sensor for the detection of protein–small molecule interactions directly in serum and other complex matrices. J. Am. Chem. Soc. 131, 6955–6957 (2009).

    Article  CAS  Google Scholar 

  17. Zuo, X. et al. A target-responsive electrochemical aptamer switch (TREAS) for reagentless detection of nanomolar ATP. J. Am. Chem. Soc. 129, 1042–1043 (2007).

    Article  CAS  Google Scholar 

  18. Das, J., Aziz, M. A. & Yang, H. A nanocatalyst-based assay for proteins: DNA-free ultrasensitive electrochemical detection using catalytic reduction of p-nitrophenol by gold-nanoparticle labels. J. Am. Chem. Soc. 128, 16022–16023 (2006).

    Article  CAS  Google Scholar 

  19. Das, J., Jo, K., Lee, J. W. & Yang, H. Electrochemical immunosensor using p-aminophenol redox cycling by hydrazine combined with a low background current. Anal. Chem. 79, 2790–2796 (2007).

    Article  CAS  Google Scholar 

  20. Das, J., Lee, J-A. & Yang, H. Ultrasensitive detection of DNA in diluted serum using NaBH4 electrooxidation mediated by [Ru(NH3)6)]3+ at indium–tin oxide electrodes. Langmuir 26, 6804–6808 (2010).

    Article  CAS  Google Scholar 

  21. Xiang, Y., Xie, M., Bash, R., Chen, J. J. L. & Wang, J. Ultrasensitive label-free aptamer-based electronic detection. Angew. Chem. Int. Ed. 119, 9212–9214 (2007).

    Article  Google Scholar 

  22. Plaxco, K. W. & Soh, H. T. Switch-based biosensors: a new approach towards real-time, in vivo molecular detection. Trends Biotechnol. 29, 1–5 (2011).

    Article  CAS  Google Scholar 

  23. Lai, R. L. et al. Rapid, sequence-specific detection of unpurified PCR amplicons via a reusable, electrochemical sensor. Proc. Natl Acad. Sci. USA 103, 4017–4021 (2006).

    Article  CAS  Google Scholar 

  24. Lapierre, M. A., O'Keefe, M. M., Taft, B. J. & Kelley, S. O. Electrocatalytic detection of pathogenic DNA sequences and antibiotic resistance markers. Anal. Chem. 75, 6327–6333 (2003).

    Article  CAS  Google Scholar 

  25. Soleymani, L., Fang, Z., Sargent, E. H. & Kelley, S. O. Programming the detection limits of biosensors through controlled nanostructuring. Nature Nanotechnol. 4, 844–848 (2009).

    Article  CAS  Google Scholar 

  26. Soleymani, L. et al. Nanostructuring of patterned microelectrodes to enhance the sensitivity of electrochemical nucleic acids detection. Angew. Chem. Int. Ed. 48, 8457–8460 (2009).

    Article  CAS  Google Scholar 

  27. Yang, H. et al. Direct, electronic microRNA detection reveals differential expression profiles in 30 minutes. Angew. Chem. Int. Ed. 48, 8461–8464 (2009).

    Article  CAS  Google Scholar 

  28. Fang, Z. et al. Direct profiling of cancer biomarkers in tumour tissue using a multiplexed nanostructured microelectrode integrated circuit. ACS Nano 3, 3207–3213 (2009).

    Article  CAS  Google Scholar 

  29. Das, J. & Kelley, S. O. Protein detection using arrayed microsensor chips: tuning sensor footprint to achieve ultrasensitive readout of CA-125 in serum and whole blood. Anal. Chem. 83, 1167–1172 (2011).

    Article  CAS  Google Scholar 

  30. Soleymani, L. et al. Hierarchical nanotextured microelectrodes overcome the molecular transport barrier to achieve rapid, direct bacterial detection. ACS Nano 5, 3360–3366 (2011).

    Article  CAS  Google Scholar 

  31. Vasilyeva, E., Fang, Z., Minden, M., Sargent, E. H. & Kelley, S. O. Direct genetic analysis of ten cancer cells: tuning molecular probe design for efficient mRNA capture Angew. Chem. Int. Ed. 50, 4137–4141 (2011).

    Article  CAS  Google Scholar 

  32. Su, L., Sankar, C. G., Sen, D. & Yu, H-Z. Kinetics of ion-exchange binding of redox metal cations to thiolate–DNA monolayers on gold. Anal. Chem. 76, 5953–5959 (2004).

    Article  CAS  Google Scholar 

  33. Bernstein, J. A., Khodursky, A. B., Lin, P. H., Lin-Chao, S. & Cohen, S. N. Global analysis of mRNA decay and abundance in Escherichia coli at single-gene resolution using two-color fluorescent DNA microarrays Proc. Natl Acad. Sci. USA 99, 9697–9702 (2002).

    Article  CAS  Google Scholar 

  34. Lam, B., Fang, Z., Sargent, E. H. & Kelley, S. O. Polymerase chain reaction-free, sample-to-answer bacterial detection in 30 minutes with integrated cell lysis. Anal. Chem. 84, 21–25 (2012).

    Article  CAS  Google Scholar 

  35. Taft, B. J., O'Keefe, M., Fourkas, J. T. & Kelley, S. O. Engineering DNA-electrode connectivities: manipulation of linker length and structure. Anal. Chim. Acta 496, 81–91 (2003).

    Article  CAS  Google Scholar 

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Acknowledgements

We acknowledge the Natural Sciences and Engineering Research Council (Discovery Grant to S.O.K.), Mitacs (Elevate Fellowship to J.D.) and the Defense Advanced Research Projects Agency (DXoD programme funding to S.O.K. and E.H.S.) for their support of this work.

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Authors and Affiliations

Authors

Contributions

S.O.K. and J.D. conceived the experiments. J.D., K.B.C., A.Z. and P.L. performed the experiments. J.D., K.B.C., A.Z., P.L., E.H.S. and S.O.K. discussed the results. J.D., K.B.C., E.H.S. and S.O.K. co-wrote and edited the manuscript.

Corresponding author

Correspondence to Shana O. Kelley.

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The authors declare no competing financial interests.

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Das, J., Cederquist, K., Zaragoza, A. et al. An ultrasensitive universal detector based on neutralizer displacement. Nature Chem 4, 642–648 (2012). https://doi.org/10.1038/nchem.1367

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