Ultrafast Plasmonic Nucleic Acid Amplification and Real-Time Quantification for Decentralized Molecular Diagnostics

Abstract

Point-of-care real-time reverse-transcription polymerase chain reaction (RT-PCR) facilitates the widespread use of rapid, accurate, and cost-effective near-patient testing that is available to the public. Here, we report ultrafast plasmonic nucleic acid amplification and real-time quantification for decentralized molecular diagnostics. The plasmonic real-time RT-PCR system features an ultrafast plasmonic thermocycler (PTC), a disposable plastic-on-metal (PoM) cartridge, and an ultrathin microlens array fluorescence (MAF) microscope. The PTC provides ultrafast photothermal cycling under white-light-emitting diode illumination and precise temperature monitoring with an integrated resistance temperature detector. The PoM thin film cartridge allows rapid heat transfer as well as complete light blocking from the photothermal excitation source, resulting in real-time and highly efficient PCR quantification. Besides, the MAF microscope exhibits close-up and high-contrast fluorescence microscopic imaging. All of the systems were fully packaged in a palm size for point-of-care testing. The real-time RT-PCR system demonstrates the rapid diagnosis of coronavirus disease-19 RNA virus within 10 min and yields 95.6% of amplification efficiency, 96.6% of classification accuracy for preoperational test, and 91% of total percent agreement for clinical diagnostic test. The ultrafast and compact PCR system can decentralize point-of-care molecular diagnostic testing in primary care and developing countries.

Single-shot multi-channel plasmonic real-time polymerase chain reaction for multi-target point-of-care testing

“…The multi-color MAF images were obtained and then processed to form a single high-contrast image using a series of image reconstruction algorithms, such as array image slicing, image averaging, image subtracting, and image masking. 27 Each array fluorescence image was sliced into single images in two groups (6 single images for one color), which were then stacked and merged using Chasys Draw IES (ver. 5.23.01) image averaging tools.…”

“…21,26 More recently, handheld plasmonic RT-qPCR systems offer a rational solution for integrating plasmonic nanostructures with a thin-film temperature detector as well as developing plasticon-metal configuration for light blocking of photothermal excitation from a white light-emitting diode (WLED). 27 However, additional issues remain, which hinder the improvement of the diagnostic performance and the multicolor real-time quantification at the point-of-care level. 28 Microscopic fluorescence imaging is required to extract the exponential increase in fluorescence intensity for microfluidic PCR applications.…”

“…36 Microlens array configurations allow close-up fluorescence imaging and real-time quantification during rapid plasmonic PCR at the handheld level but still require simultaneous multi-color quantification in the small RT-qPCR system. 27,37,38 Here, we report a single-shot multi-channel plasmonic RT-qPCR (mpRT-qPCR) assay with ultrafast nanostructure-driven amplification and multi-target quantification. The mpRT-qPCR system incorporates three main steps of WLED-driven rapid photothermal heating from a plasmonic thermocycler (PTC), rapid cooling through efficient heat dissipation of an aluminum (Al) thin-film layer on a cartridge, and multi-color microlens array fluorescence (MAF) imaging (Fig.…”

Highly Efficient On-Chip Photothermal Cell Lysis for Nucleic Acid Extraction Using Localized Plasmonic Heating of Strongly Absorbing Au Nanoislands

“…Photothermal heating is actively utilized for diverse theragnostic applications such as cancer therapy, , pathogens inactivation, − and nucleic acid amplification, − including cell lysis. Photothermal heating exhibits attractive benefits such as ultrafast heating and cooling, local heat management, and specific target constituent damage using assorted light-absorbing materials such as metallic thin films or nanoparticles (NPs). , In particular, Au-coated polycarbonate membrane rapidly ruptures the membrane proteins of infectious pathogens for point-of-care urinalysis .…”

Concave microlens arrays with tunable curvature for enhanced photodegradation of organic pollutants in water: A non‐contact approach

“…4,5 Microlenses (MLs) have emerged as a promising technology to enhance energy transfer efficiency by optimizing light inside of the solar-driven reactors. [6][7][8][9] The near-field effect of MLs enables the strong focusing effects in a short distance around the MLs, 10,11 which has been widely applied in the fields where high light harvesting efficiency [12][13][14][15][16][17] and minimized dimensions [18][19][20] are required. The focal distance of typical convex MLs is in the scale of micrometers, 11,21,22 so they have to be implemented inside of a reactor to make focal points located inside water.…”