Abstract
Current visual biosensing methods, including colorimetric-based, fluorescence-based and chemiluminescence-based methods, are inappropriate for the hundreds of millions of people affected by color blindness and color weakness. Compared with these available methods, a droplet motion-based strategy might be a promising protocol for extension to a wider user base. Here we report a protocol for manipulating the hydrophobicity of DNA, which offers a droplet motion-based biosensing platform for the visual detection of small molecules (ATP), nucleic acids (microRNA) and proteins (thrombin). The protocol starts with target-triggered rolling-circle amplification that can readily generate short single-stranded DNA (ssDNA) fragments or long ssDNA. By exploiting macroscopic wetting behavior and molecular interaction, one can tailor the conformation of ssDNA on the water–oil interface to control the relevant DNA hydrophobicity. The wettability of DNA can be translated into visual signals via reading the sliding speed or the critical sliding angle. The time range for the entire protocol is ∼1 d, and the detection process takes ∼1 min.
This is a preview of subscription content, access via your institution
Access options
Access Nature and 54 other Nature Portfolio journals
Get Nature+, our best-value online-access subscription
$29.99 / 30 days
cancel any time
Subscribe to this journal
Receive 12 print issues and online access
$259.00 per year
only $21.58 per issue
Buy this article
- Purchase on Springer Link
- Instant access to full article PDF
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
Data availability
The authors declare that the main data supporting the findings of this study are available within the article and its Supplementary Information files.
References
Sawyers, C. L. The cancer biomarker problem. Nature 452, 548–552 (2008).
Bonventre, J. V., Vaidya, V. S., Schmouder, R., Feig, P. & Dieterle, F. Next-generation biomarkers for detecting kidney toxicity. Nat. Biotechnol. 28, 436–440 (2010).
Schwarzenbach, H., Hoon, D. S. B. & Pantel, K. Cell-free nucleic acids as biomarkers in cancer patients. Nat. Rev. Cancer 11, 426–437 (2011).
Ferrari, M. Cancer nanotechnology: opportunities and challenges. Nat. Rev. Cancer 5, 161–171 (2005).
Lequin, R. M. Enzyme immunoassay (EIA)/enzyme-linked immunosorbent assay (ELISA). Clin. Chem. 51, 2415–2418 (2005).
Li, H. X. & Rothberg, L. J. Label-free colorimetric detection of specific sequences in genomic DNA amplified by the polymerase chain reaction. J. Am. Chem. Soc. 126, 10958–10961 (2004).
Laing, S., Jamieson, L. E., Faulds, K. & Graham, D. Surface-enhanced Raman spectroscopy for in vivo biosensing. Nat. Rev. Chem. 1, 0060 (2017).
Wang, M., Wang, C. Y. & Han, X. L. Selection of internal standards for accurate quantification of complex lipid species in biological extracts by electrospray ionization mass spectrometry—what, how and why? Mass Spectrom. Rev. 36, 693–714 (2017).
Rosi, N. L. & Mirkin, C. A. Nanostructures in biodiagnostics. Chem. Rev. 105, 1547–1562 (2005).
Lei, J. P. & Ju, H. X. Signal amplification using functional nanomaterials for biosensing. Chem. Soc. Rev. 41, 2122–2134 (2012).
Kelley, S. O. et al. Advancing the speed, sensitivity and accuracy of biomolecular detection using multi-length-scale engineering. Nat. Nanotechnol. 9, 969–980 (2014).
Wang, J. Electrochemical biosensors: towards point-of-care cancer diagnostics. Biosens. Bioelectron. 21, 1887–1892 (2006).
Lu, L. M., Zhang, X. B., Kong, R. M., Yang, B. & Tan, W. A ligation-triggered dnazyme cascade for amplified fluorescence detection of biological small molecules with zero-background signal. J. Am. Chem. Soc. 133, 11686–11691 (2011).
Xiang, Y. & Lu, Y. Using personal glucose meters and functional DNA sensors to quantify a variety of analytical targets. Nat. Chem. 3, 697–703 (2011).
Quesada-Gonzalez, D. & Merkoci, A. Nanomaterial-based devices for point-of-care diagnostic applications. Chem. Soc. Rev. 47, 4697–4709 (2018).
Yan, J. T., Lee, S., Zhang, A. F. & Yoon, J. Self-immolative colorimetric, fluorescent and chemiluminescent chemosensors. Chem. Soc. Rev. 47, 6900–6916 (2018).
Fu, E. Enabling robust quantitative readout in an equipment-free model of device development. Analyst 139, 4750–4757 (2014).
Tian, T. et al. Distance-based microfluidic quantitative detection methods for point-of-care testing. Lab Chip 16, 1139–1151 (2016).
Gao, Z. F. et al. Naked-eye point-of-care testing platform based on a pH-responsive superwetting surface: toward the non-invasive detection of glucose. NPG Asia Mater. 10, 177–189 (2018).
Chen, Y. et al. Double-enzymes-mediated bioluminescent sensor for quantitative and ultrasensitive point-of-care testing. Anal. Chem. 89, 5422–5427 (2017).
Martinez, A. W., Phillips, S. T., Butte, M. J. & Whitesides, G. M. Patterned paper as a platform for inexpensive, low-volume, portable bioassays. Angew. Chem. Int. Ed. 46, 1318–1320 (2007).
Willner, I. & Zayats, M. Electronic aptamer-based sensors. Angew. Chem. Int. Ed. 46, 6408–6418 (2007).
Lin, M. H. et al. Programmable engineering of a biosensing interface with tetrahedral DNA nanostructures for ultrasensitive DNA detection. Angew. Chem. Int. Ed. 54, 2151–2155 (2015).
Monserud, J. H. & Schwartz, D. K. Effects of molecular size and surface hydrophobicity on oligonucleotide interfacial dynamics. Biomacromolecules 13, 4002–4011 (2012).
Elder, R. M. & Jayaraman, A. Structure and thermodynamics of ssDNA oligomers near hydrophobic and hydrophilic surfaces. Soft Matter 9, 11521–11533 (2013).
Pei, H., Zuo, X., Zhu, D., Huang, Q. & Fan, C. Functional DNA nanostructures for theranostic applications. Acc. Chem. Res. 47, 550–559 (2014).
Zhan, S. S., Pan, Y., Gao, Z. F., Lou, X. D. & Xia, F. Biological and chemical sensing applications based on special wettable surfaces. TrAC Trend Anal. Chem. 108, 183–194 (2018).
Gao, Z. F. et al. Controlling droplet motion on an organogel surface by tuning the chain length of DNA and its biosensing application. Chem 4, 2929–2943 (2018).
Deng, X., Mammen, L., Butt, H. J. & Vollmer, D. Candle soot as a template for a transparent robust superamphiphobic coating. Science 335, 67–70 (2012).
Tian, X. L., Verho, T. & Ras, R. H. A. Moving superhydrophobic surfaces toward real-world applications. Science 352, 142–143 (2016).
Shang, L. R., Cheng, Y. & Zhao, Y. J. Emerging droplet microfluidics. Chem. Rev. 117, 7964–8040 (2017).
Xu, T. L. et al. Superwettable electrochemical biosensor toward detection of cancer biomarkers. ACS Sens. 3, 72–78 (2018).
Liu, X. J. et al. 3D printing of bioinspired liquid superrepellent structures. Adv. Mater. 30, 1800103 (2018).
Feng, W. Q., Ueda, E. & Levkin, P. A. Droplet microarrays: from surface patterning to high-throughput applications. Adv. Mater. 30, 1706111 (2018).
Qin, M. et al. A rainbow structural-color chip for multisaccharide recognition. Angew. Chem. Int. Ed. 55, 6911–6914 (2016).
Chi, J. J. et al. Patterned photonic nitrocellulose for pseudopaper ELISA. Anal. Chem. 89, 7727–7733 (2017).
Lei, Y. J., Sun, R. Z., Zhang, X. C., Feng, X. J. & Jiang, L. Oxygen-rich enzyme biosensor based on superhydrophobic electrode. Adv. Mater. 28, 1477–1481 (2016).
Xu, L. P. et al. Ultratrace DNA detection based on the condensing-enrichment effect of superwettable microchips. Adv. Mater. 27, 6878–6884 (2015).
Yang, S. K., Dai, X. M., Stogin, B. B. & Wong, T. S. Ultrasensitive surface-enhanced Raman scattering detection in common fluids. Proc. Natl Acad. Sci. USA 113, 268–273 (2016).
Wong, T. S. et al. Bioinspired self-repairing slippery surfaces with pressure-stable omniphobicity. Nature 477, 443–447 (2011).
Leslie, D. C. et al. A bioinspired omniphobic surface coating on medical devices prevents thrombosis and biofouling. Nat. Biotechnol. 32, 1134–1140 (2014).
Howell, C., Grinthal, A., Sunny, S., Aizenberg, M. & Aizenberg, J. Designing liquid-infused surfaces for medical applications: a review. Adv. Mater. 30, e1802724 (2018).
He, W., Liu, P., Zhang, J. & Yao, X. Emerging applications of bioinspired slippery surfaces in biomedical fields. Chem. Eur. J. 24, 14864–14877 (2018).
Liu, P. et al. Condensation-assisted micro-patterning of low-surface-tension liquids on reactive oil-repellent surfaces. J. Mater. Chem. A 5, 16344–16351 (2017).
Lv, J. Y., Yao, X., Zheng, Y. M., Wang, J. J. & Jiang, L. Antiadhesion organogel materials: from liquid to solid. Adv. Mater. 29, 1703032 (2017).
Liu, H. L., Zhang, P. C., Liu, M. J., Wang, S. T. & Jiang, L. Organogel-based thin films for self-cleaning on various surfaces. Adv. Mater. 25, 4477–4481 (2013).
Yao, X., Ju, J., Yang, S., Wang, J. J. & Jiang, L. Temperature-driven switching of water adhesion on organogel surface. Adv. Mater. 26, 1895–1900 (2014).
Yao, X. et al. Self-replenishable anti-waxing organogel materials. Angew. Chem. Int. Ed. 54, 8975–8979 (2015).
Liu, M. J., Xue, Z. X., Liu, H. & Jiang, L. Surface wetting in liquid–liquid–solid triphase systems: solid-phase-independent transition at the liquid-liquid interface by lewis acid–base interactions. Angew. Chem. Int. Ed. 51, 8348–8351 (2012).
Guo, T. Q., Che, P. D., Heng, L. P., Fan, L. Z. & Jiang, L. Anisotropic slippery surfaces: electric-driven smart control of a drop’s slide. Adv. Mater. 28, 6999–7007 (2016).
Li, X. et al. Cascaded signal amplification via target-triggered formation of aptazyme for sensitive electrochemical detection of ATP. Biosens. Bioelectron. 102, 296–300 (2018).
Gong, X. et al. Target recycling amplification for label-free and sensitive colorimetric detection of adenosine triphosphate based on un-modified aptamers and DNAzymes. Anal. Chim. Acta 828, 80–84 (2014).
Li, X., Peng, Y., Chai, Y. Q., Yuan, R. & Xiang, Y. A target responsive aptamer machine for label-free and sensitive non-enzymatic recycling amplification detection of ATP. Chem. Commun. 52, 3673–3676 (2016).
Huang, Y., Lei, J. P., Cheng, Y. & Ju, H. X. Target-assistant Zn2+-dependent DNAzyme for signal-on electrochemiluminescent biosensing. Electrochim. Acta 155, 341–347 (2015).
Gao, Z. F. et al. Guanine nanowire based amplification strategy: enzyme-free biosensing of nucleic acids and proteins. Biosens. Bioelectron. 78, 351–357 (2016).
Qavi, A. J., Kindt, J. T., Gleeson, M. A. & Bailey, R. C. Anti-DNA: RNA antibodies and silicon photonic microring resonators: increased sensitivity for multiplexed microRNA detection. Anal. Chem. 83, 5949–5956 (2011).
Ou, X. et al. Simultaneous detection of telomerase and miRNA with graphene oxide-based fluorescent aptasensor in living cells and tissue samples. Biosens. Bioelectron. 124, 199–204 (2019).
Sipova, H. et al. Surface plasmon resonance biosensor for rapid label-free detection of microribonucleic acid at subfemtomole level. Anal. Chem. 82, 10110–10115 (2010).
Xiao, Y., Lubin, A. A., Heeger, A. J. & Plaxco, K. W. Label-free electronic detection of thrombin in blood serum by using an aptamer-based sensor. Angew. Chem. Inter. Ed. 44, 5456–5459 (2005).
Bini, A., Minunni, M., Tombelli, S., Centi, S. & Mascini, M. Analytical performances of aptamer-based sensing for thrombin detection. Anal. Chem. 79, 3016–3019 (2007).
Mani, R. J., Dye, R. G., Snider, T. A., Wang, S. & Clinkenbeard, K. D. Bi-cell surface plasmon resonance detection of aptamer mediated thrombin capture in serum. Biosens. Bioelectron. 26, 4832–4836 (2011).
Wang, Y. & Liu, B. Conjugated polyelectrolyte-sensitized fluorescent detection of thrombin in blood serum using aptamer-immobilized silica nanoparticles as the platform. Langmuir 25, 12787–12793 (2009).
Simon, A. J., Vallee-Belisle, A., Ricci, F. & Plaxco, K. W. Intrinsic disorder as a generalizable strategy for the rational design of highly responsive, allosterically cooperative receptors. Proc. Natl Acad. Sci. USA 111, 15048–15053 (2014).
Dean, J. A. Lange’s Handbook of Chemistry (McGraw-Hill Professional, 2017).
Acknowledgements
This work was supported by the National Natural Science Foundation of China (21535002) to S.Z.; the National Natural Science Foundation of China (21525523, 21722507, 21574048 and 21874121), the National Basic Research Program of China (973 Program, 2015CB932600) and the National Key R&D Program of China (2017YFA0208000) to F.X.; the National Natural Science Foundation of China (31800829), the Natural Science Foundation of Shandong Province (ZR2018BB054) and PhD Research Foundation of Linyi University (LYDX2018BS005) to Z.F.G.; and the National Natural Science Foundation of China (21703210) and the China Postdoctoral Science Foundation (2017M610492) to R.L.
Author information
Authors and Affiliations
Contributions
Z.F.G., F.X. and L.J. designed the research. Z.F.G., R.L., J.W. and J.D. performed the research. J.D., W.-H.H., M.L. and S.Z. contributed new reagents and analytic tools. Z.F.G., R.L., J.W., S.W., F.X., S.Z. and L.J. analyzed the data. L.J. supervised the project. Z.F.G., R.L., F.X. and S.Z. wrote the paper.
Corresponding authors
Ethics declarations
Competing interests
The authors declare no competing interests.
Additional information
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Related links
Key references using this protocol
Gao, Z. F. et al. Chem 4, 2929–2943 (2018): https://doi.org/10.1016/j.chempr.2018.09.028
Gao, Z. F. et al. NPG Asia Mater. 10, 177–189 (2018): https://doi.org/10.1038/s41427-018-0024-7
Zhan, S., Pan, Y., Gao, Z. F., Lou, X. D. & Xia, F. TrAC Trends Anal. Chem. 108, 183–194 (2018): https://doi.org/10.1016/j.trac.2018.09.001
Integrated supplementary information
Supplementary Fig. 1. Gel electrophoresis image after RCA.
(1) DNA size marker; (2) padlock probe; (3) padlock probe circularization with ATP; (4) RCA reaction with ATP; (5) padlock probe circularization without ATP; (6) RCA reaction without ATP. The ATP concentration is 500 μM. Adapted with permission from ref. 28, Cell Press.
Supplementary Fig. 2. Control experiment and practical performance of ATP detection.
(a) The recorded surface tensions (black) and CSAs (red) at various ATP concentrations. The CSA and surface tension of pure ATP solutions barely changed with varying concentrations of ATP. Error bars represent the standard deviation from at least six individual measurements. Arrows refer to the black and red data corresponding to the black and red axes on the left and right, respectively. (b) Response of CSAs to 0 nM, 0.5 nM, 50 nM, and 5000 nM ATP in buffer and human serum solution, respectively. A standard addition method was next conducted in spiked human serum samples (diluted 200-fold before use) to verify the accuracy of the system55. Based on the highly specific binding of ATP to the T4 DNA ligase, the CSA recorded in human serum was comparable to that in buffer solution at all ATP concentrations, indicating this protocol could be effectively applied in complicated samples. The control experiments, diluted serum sample and buffer solution without ATP, proved that the serum sample containing sugars, proteins, lipids, etc., has a negligible effect on the CSA. Error bars represent the standard deviation from at least six individual measurements. Adapted with permission from ref. 28, Cell Press.
Supplementary Fig. 3. Control experiments for miR-21 and thrombin detection.
The recorded surface tensions (black) and CSAs (red) at various (a) miR-21, (b) thrombin concentrations. The CSA and surface tension barely changed with varying concentrations of miRNA or thrombin. Error bars represent the standard deviation from at least six individual measurements. Arrows refer to the black and red data corresponding to the black and red axes on the left and right, respectively.
Supplementary information
Supplementary Information
Supplementary Figs. 1–3 and Supplementary Table 1
Supplementary Video 1
The distribution of the water–oil interface between the RCA droplet without ATP and the organogel surface, which was studied by an inverted fluorescence microscope tilted from 0° to 8.1°. The bottom-view video was recorded in the grayscale oil channel. The movie playback accelerated ten times. Adapted with permission from ref. 1, Cell Press. This movie shows that the RCA droplet without ATP barely moved on the organogel surface because the oil layer was damaged due to the strong hydrophobic interaction between short DNA fragments and oil molecules.
Supplementary Video 2
The distribution of the water–oil interface between the RCA droplet with ATP and the organogel surface, which was studied by an inverted fluorescence microscope tilted from 0° to 8.1°. The bottom-view video is recorded at grayscale oil channel. The shifts at ~60 s and 105 s result from camera movement. The movie playback is accelerated ten times. This movie shows the RCA droplet with ATP sliding easily on the organogel surface because the oil layer was almost intact due to the weak hydrophobic interaction between long ssDNA and oil molecules. Adapted with permission from ref. 1, Cell Press.
Supplementary Video 3
Sliding behavior of RCA droplets with ATP on the organogel surface. The droplet is stained with methylene blue to display clearly. The droplet size is 2 μL. Adapted with permission from ref. 1, Cell Press.
Supplementary Video 4
Sliding behavior of RCA droplets without ATP on the organogel surface. The droplet is stained with rhodamine B to display clearly. The droplet size is 2 μL. The movie playback is accelerated eight times. Adapted with permission from ref. 1, Cell Press.
Rights and permissions
About this article
Cite this article
Gao, Z.F., Liu, R., Wang, J. et al. Manipulating the hydrophobicity of DNA as a universal strategy for visual biosensing. Nat Protoc 15, 316–337 (2020). https://doi.org/10.1038/s41596-019-0235-6
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/s41596-019-0235-6
This article is cited by
-
Superwettable interface towards biodetection in confined space
Nano Research (2024)
Comments
By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.