Digital Resolution Detection Without Droplets or Enzymes through Photonic Crystal Amplification
Brian Cunningham,
Donald Biggar Willett Professor in Engineering,
University of Illinois at Urbana-Champaign
Droplet digital biomarker detection technologies achieve ultrasensitive detection limits by partitioning the test sample into thousands of small volumes that confine the fluorescent products of an enzymatic amplification reaction, followed by counting bright droplets. Using photonic crystal surfaces to amplify the fluorescence of light-emitting tags such as quantum dots and plasmonic fluors, individual nucleic acid biomarkers, proteins, and virus particles can be digitally counted without sample partitioning or enzymatic reactions, resulting in simple workflow, rapid sample-to-answer time, and ultrasensitivity using simple inexpensive instruments. We have also developed digital alternatives to PCR that offer room temperature, isothermal biochemistry methods based upon nucleic acid strand displacement reactions and CRISPR/Cas technology which rapidly amplify single target nucleic acid biomarkers (miRNA and ctDNA) into large numbers of gold nanoparticles that are digitally counted by Photonic Resonator Absorption Microscopy (PRAM). We show that an “amplify-then-digitize” strategy results in detection limits below PCR while utilizing a simple protocol and inexpensive instrument. Overall, we envision application of the biodetection technologies and assay methods towards ultrasensitive and multiplexed point of care diagnostics.
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