Plasmonic Nanoparticle Conjugation for Nucleic Acid Biosensing.

This chapter details the use of gold nanorods conjugated with peptide nucleic acid probes for sequence-specific detection of circulating tumor DNA (ctDNA). ctDNA is gaining increased attention as a biomarker for liquid biopsy, the process of detecting molecules in the peripheral blood rather than a tissue sample. It has wide ranging applications as a diagnostic and prognostic biomarker with a similar mutational profile as the tumor. Plasmonic nanoparticles offer a relatively rapid, amplification-free method for detection of ctDNA through the use of sequence-specific peptide nucleic acid (PNA) probes. In this chapter, we discuss methods for probe design, conjugation to plasmonic particles, and ctDNA quantitation with the resulting sensor. This chapter is a resource for those looking to use plasmonic gold particles for sensing in a solution format for a range of applications.

Vibrant reflective sensors with percolation film Fabry-Pérot nanocavities.

Dynamically reconfigurable structural colors are promising materials for new smart optical systems. However, improved reflected color quality (e.g., saturation, optical contrast, angular invariance) and larger tuning range/sensitivity are needed. Here, we demonstrate a vibrant, actively tunable system which meets these needs via coupling broadband plasmonic resonators to a responsive polymer film. Our structure consists of near-percolation gold nanoislands deposited on a poly[methyl methacrylate] (PMMA) spacer above a gold mirror, forming a Fabry-Pérot nanocavity. Broadband absorption in this system creates vivid reflected colors, while the polymer spacer enables continuous tuning over a wide color space. By exploiting swelling effects in PMMA, we show fast, reversible color switching in response to organic vapors. Our sensitive optical structure amplifies small vapor-induced changes in the spacer thickness, enabling naked-eye detection of changes as small as 10 nm. Additionally, optical absorption >99% yields modulation contrasts up to 80:1, opening the door to ultra-sensitive on-chip signal measurements, complementing the visual colorimetric readout. This structure has immediate implications for colorimetric bio/chemical sensing and may also find application to reflective displays and flexible/adaptive optical coatings.

Rational design of on-chip gold plasmonic nanoparticles towards ctDNA screening.

This paper demonstrates the design, synthesis, simulation, and testing of three distinct geometries of plasmonic gold nanoparticles for on-chip DNA screening towards liquid biopsy. By employing a seed-mediated growth method, we have synthesized gold nanospheres, nanorods, and nanobipyramids. In parallel, we developed numerical simulations to understand the effects of nanoparticle geometry on the resonance features and refractive index sensitivity. Both experimental and simulation results were compared through a series of studies including in-solution and on-chip tests. We have thoroughly characterized the impact of nanoparticle geometry on the sensitivity to circulating tumor DNA, with immediate implications for liquid biopsy. The results agree well with theoretical predictions and simulations, including both bulk refractive index sensitivity and thin film sensitivity. Importantly, this work quantitatively establishes the link between nanoparticle geometry and efficacy in detecting rare circulating biomarkers. The nanobipyramids provided the highest sensitivity, approximately doubling the sensitivity compared to nanorods. To the best of our knowledge this is the first report carrying through geometric effects of simulation to clinically relevant biosensing. We put forth here synthesis and testing of three nanoparticle geometries, and a framework for both experimental and theoretical validation of plasmonic sensitivities towards liquid biopsy.

Scalable Signature-Based Molecular Diagnostics Through On-chip Biomarker Profiling Coupled with Machine Learning.

Molecular diagnostics have traditionally relied on discrete biological substances as diagnostic markers. In recent years however, advances in on-chip biomarker screening technologies and data analytics have enabled signature-based diagnostics. Such diagnostics aim to utilize unique combinations of multiple biomarkers or diagnostic 'fingerprints' rather than discrete analyte measurements. This approach has shown to improve both diagnostic accuracy and diagnostic specificity. In this review, signature-based diagnostics enabled by microfluidic and micro-/nano- technologies will be reviewed with a focus on device design and data analysis pipelines and methodologies. With increasing amounts of data available from microfluidic biomarker screening, isolation, and detection platforms, advanced data handling and analytics approaches can be employed. Thus, current data analysis approaches including machine learning and recent advances with image processing, along with potential future directions will be explored. Lastly, the needs and gaps in current literature will be elucidated to inform future efforts towards development of molecular diagnostics and biomarker screening technologies.

Scalable COVID-19 Detection Enabled by Lab-on-Chip Biosensors.

Introduction: The emergence of a novel coronavirus, SARS-CoV-2, has highlighted the need for rapid, accurate, and point-of-care diagnostic testing. As of now, there is not enough testing capacity in the world to meet the stated testing targets, which are expected to skyrocket globally for broader testing during reopening. Aim: This review focuses on the development of lab-on-chip biosensing platforms for diagnosis of COVID-19 infection. Results: We discuss advantages of utilizing lab-on-chip technologies in response to the current global pandemic, including their potential for low-cost, rapid sample-to-answer processing times, and ease of integration into a range of healthcare settings. We then highlight the development of magnetic, colorimetric, plasmonic, electrical, and lateral flow-based lab-on-chip technologies for the detection of SARS-CoV-2, in addition to other viruses. We focus on rapid, point-of-care technologies that can be deployed at scale, as such devices could be promising alternatives to the current gold standard of reverse transcription-polymerase chain reaction (RT-PCR) diagnostic testing. Conclusion: This review is intended to provide an overview of the current state-of-the-field and serve as a resource for innovative development of new lab-on-chip assays for COVID-19 detection.

Advances in diagnostic microfluidics.

Microfluidics is an emerging field in diagnostics that allows for extremely precise fluid control and manipulation, enabling rapid and high-throughput sample processing in integrated micro-scale medical systems. These platforms are well-suited for both standard clinical settings and point-of-care applications. The unique features of microfluidics-based platforms make them attractive for early disease diagnosis and real-time monitoring of the disease and therapeutic efficacy. In this chapter, we will first provide a background on microfluidic fundamentals, microfluidic fabrication technologies, microfluidic reactors, and microfluidic total-analysis-systems. Next, we will move into a discussion on the clinical applications of existing and emerging microfluidic platforms for blood analysis, and for diagnosis and monitoring of cancer and infectious disease. Together, this chapter should elucidate the potential that microfluidic systems have in the development of effective diagnostic technologies through a review of existing technologies and promising directions.

Metallic photonic crystal-based sensor for cryogenic environments.

We investigate the design, characterization, and application of metallic photonic crystal (MPC) structures, consisting of plasmonic gold nanogratings on top of a photonic waveguide, as transducers for lab-on-chip biosensing in cryogenic environments. The compact design offers a promising approach to sensitive, in situ biosensing platforms for astrobiology applications (e.g., on the "icy moons" of the outer solar system). We fabricated and experimentally characterized three MPC sensor geometries, with variable nanograting width, at temperatures ranging from 300 K to 180 K. Sensors with wider nanogratings were more sensitive to changes in the local dielectric environment. Temperature-dependent experiments revealed an increase in plasmonic resonance intensity of around 13% at 180 K (compared with 300 K), while the coupled plasmonic-photonic resonance was less sensitive to temperature, varying by less than 5%. Simulation results confirm the relative temperature stability of the plasmonic-photonic mode and, combined with its high sensitivity, suggest a novel application of this mode as the sensing transduction mechanism over wide temperature ranges. To our knowledge, this is among the first reports of the design and characterization of a nanoplasmonic sensor specifically for low-temperature sensing operation.

Design of peptide nucleic acid probes on plasmonic gold nanorods for detection of circulating tumor DNA point mutations.

Here we present a gold nanorod-based platform for the sequence-specific detection of circulating tumor DNA (ctDNA) point mutations without the need for amplification or fluorescence labeling. Peptide nucleic acid probes complimentary to the G12V mutation in the KRAS gene were conjugated to gold nanorods, and the localized surface plasmon resonance absorbance through the sample was measured after exposure to synthetic ctDNA at various concentrations. Each step of the reaction was thoroughly controlled, starting from reagent concentrations and including conjugation, sonication, and incubation time. The platform was evaluated in both buffer and spiked healthy patient serum, demonstrating a linear working range below 125 nanograms of ctDNA per milliliter solution, and an effective limit of detection of 2 nanograms of ctDNA per milliliter. A clear distinction between mutant and wild type synthetic ctDNA was also found using this platform. In order to improve upon the selectivity of the sensor, a DNA hybridization simulation was performed to understand how the addition of mutations to the peptide nucleic acid probe could enhance the selectivity for capture of mutant over wild type sequences. The top candidate from the simulations, which had an additional mutation two base pairs away from the mutation of interest, had a significant impact on the selectivity between mutant and wild type capture. This paper provides a framework for sequence-specific capture of ctDNA, and a method of improving selectivity for desired point mutations through careful probe design.

Microfluidics-enabled rapid manufacturing of hierarchical silica-magnetic microflower toward enhanced circulating tumor cell screening.

The emergence of microfluidic techniques provides new opportunities for chemical synthesis and biomedical applications. Herein, we first develop a microfluidics-based flow and sustainable strategy to synthesize hierarchical silica-magnetic microflower with unique multilayered structure for the efficient capture of circulating tumor cells through our engineered microfluidic screening chip. The production of microflower materials can be realized within 94 milliseconds and a yield of nearly 5 grams per hour can be achieved. The enhanced bioaccessibility of such a multilayered microflower towards cancer cells (MCF-7 and MDA-MB-231) is demonstrated, and the cancer cell capture efficiency of this hierarchical immunomagnetic system in clinical blood samples is significantly increased compared with a standard CellSearch™ assay. These findings bring new insights for engineering functional micro-/nanomaterials in liquid biopsy.

Advances in liquid biopsy on-chip for cancer management: Technologies, biomarkers, and clinical analysis.

Liquid biopsy, as a minimally invasive method of gleaning insight into the dynamics of diseases through a patient fluid sample, has been growing in popularity for cancer diagnosis, prognosis, and monitoring. While many technologies have been developed and validated in research laboratories, there has also been a push to expand these technologies into other clinical settings and as point of care devices. In this article, we discuss and evaluate microchip-based technologies for circulating tumor cell (CTC), exosome, and circulating tumor nucleic acid (ctNA) capture, detection, and analysis. Such integrated systems streamline otherwise multiple-step, manual operations to get a sample-to-answer quantitation. In addition, analysis of disease biomarkers is suited to point of care settings because of ease of use, low consumption of sample and reagents, and high throughput. We also cover the basics of biomarkers and their detection in biological fluid samples suitable for liquid biopsy on-chip. We focus on emerging technologies that process a small patient sample with high spatial-temporal resolution and derive clinically meaningful results through on-chip biomarker sensing and downstream molecular analysis in a simple workflow. This critical review is meant as a resource for those interested in developing technologies for capture, detection, and analysis platforms for liquid biopsy in a variety of settings.

Microfluidics-mediated self-template synthesis of anisotropic hollow ellipsoidal mesoporous silica nanomaterials.

Herein, a facile strategy was firstly developed to synthesize ellipsoidal mesoporous silica nanomaterials (MSNs) with well-ordered parallel channels along the short axis. A miniaturized microfluidic device with spiral-shaped channels was then chosen as a straightforward and general platform to produce the corresponding hollow counterparts of MSNs under mild conditions. Such reaction process carried out in a microfluidic system was further demonstrated to be more rapid and efficient than conventional batch method under equivalent experimental conditions. The evolution of hollow structure can be well-tuned by flow rates (i.e., etching time), providing new paradigm for rational design and engineering of anisotropic nanostructures.

Liquid biopsy on chip: a paradigm shift towards the understanding of cancer metastasis.

This comprehensive review serves as a guide for developing scalable and robust liquid biopsies on chip for capture, detection, and analysis of circulating tumor cells (CTCs). Liquid biopsy, the detection of biomarkers from body fluids, has proven challenging because of CTC rarity and the heterogeneity of CTCs shed from tumors. The review starts with the underlying biological mechanisms that make liquid biopsy a challenge before moving into an evaluation of current technological progress. Then, a framework for evaluation of the technologies is presented with special attention to throughput, capture rate, and cell viability for analysis. Technologies for CTC capture, detection, and analysis will be evaluated based on these criteria, with a focus on current approaches, limitations and future directions. The paper provides a critical review for microchip developers as well as clinical investigators to build upon the existing progress towards the goal of designing CTC capture, detection, and analysis platforms.