Abstract
This protocol describes a signal-generation mechanism for the naked-eye detection of analytes at low concentrations with ELISA. The key step is to generate solutions of desired tonality by growing gold nanoparticles with a particular state of aggregation. This is accomplished by linking the growth of gold nanoparticles with the biocatalytic cycle of the enzyme label. The protocol adapts a conventional ELISA procedure with catalase-labeled antibodies. The enzyme consumes hydrogen peroxide, and then gold (III) ions are added to generate gold nanoparticles. The concentration of hydrogen peroxide dictates the state of aggregation of gold nanoparticles. This allows for the naked-eye detection of analytes by observing the generation of blue- or red-colored gold nanoparticle solutions. When coupled with conventional ELISA, this signal-generation procedure allows for the naked-eye detection of analytes within 1 h.
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Acknowledgements
M.M.S. thanks the Engineering and Physical Sciences Research Council and the European Research Council starting investigator grant 'Naturale' for funding. This research was supported by a Marie Curie Intra-European Fellowship within the 7th European Community Framework Programme (R.d.l.R.).
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R.d.l.R. conceived and designed the method and wrote the manuscript. M.M.S. supervised the work and revised the manuscript.
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Supplementary Figure 1
Generation of colored gold nanoparticle solutions at different final concentrations of hydrogen peroxide. Blue solutions can be obtained when the concentration of hydrogen peroxide in the growth solution decreases by as low as 50 nM. Lower decreases in the concentration of hydrogen peroxide were not assayed because it is extremely difficult to prepare hydrogen peroxide solutions with accurate concentration between 120 μM and 119.95 μM. Therefore the decrease in the concentration of hydrogen peroxide needed to obtain blue-colored nanoparticle solutions could be even lower than 50 nM. (PDF 533 kb)
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de la Rica, R., Stevens, M. Plasmonic ELISA for the detection of analytes at ultralow concentrations with the naked eye. Nat Protoc 8, 1759–1764 (2013). https://doi.org/10.1038/nprot.2013.085
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DOI: https://doi.org/10.1038/nprot.2013.085
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