Seung-min Park

A method for nanofluidic device prototyping using elastomeric collapse

Nanofluidics represents a promising solution to problems in fields ranging from biomolecular analysis to optical property tuning. Recently a number of simple nanofluidic fabrication techniques have been introduced that exploit the deformability of elastomeric materials like polydimethylsiloxane (PDMS). These techniques are limited by the complexity of the devices that can be fabricated, which can only create straight or irregular channels normal to the direction of an applied strain. Here, we report a technique for nanofluidic fabrication based on the controlled collapse of microchannel structures. As is demonstrated, this method converts the easy to control vertical dimension of a PDMS mold to the lateral dimension of a nanochannel. We demonstrate here the creation of complex nanochannel structures as small as 60 nm and provide simple design rules for determining the conditions under which nanochannel formation will occur. The applicability of the technique to biomolecular analysis is demonstrated by showing DNA elongation in a nanochannel and a technique for optofluidic surface enhanced Raman detection of nucleic acids.

Selection and elution of aptamers using nanoporous sol-gel arrays with integrated microheaters

RNA and DNA aptamers that bind to target molecules with high specificity and affinity have been a focus of diagnostics and therapeutic research. These aptamers are obtained by SELEX (Systematic Evolution of Ligands by EXponential enrichment) often requiring more than 10 successive cycles of selection and amplification, where each cycle normally takes 2 days per cycle of SELEX. Here, we have demonstrated the use of sol-gel arrays of proteins in a microfluidic system for efficient selection of RNA aptamers against multiple target molecules. The microfluidic chip incorporates five sol-gel binding droplets, within which specific target proteins are imbedded. The droplets are patterned on top of individually addressable electrical microheaters used for selective elution of aptamers bound to target proteins in the sol-gel droplets. We demonstrate that specific aptamers bind their respective protein targets and can be selectively eluted by micro-heating. Finally, our microfluidic SELEX system greatly improved selection efficiency, reducing the number of selection cycles needed to produce high affinity aptamers. The process is readily scalable to larger arrays of sol-gel-embedded proteins. To our knowledge, this is the first demonstration of a chip-based selection of aptamers using microfluidics, thereby allowing development of a high throughput and efficient SELEX procedures.

Towards clinically translatable in vivo nanodiagnostics

Nanodiagnostics as a field makes use of fundamental advances in nanobiotechnology to diagnose, characterize and manage disease at the molecular scale. As these strategies move closer to routine clinical use, a proper understanding of different imaging modalities, relevant biological systems and physical properties governing nanoscale interactions is necessary to rationally engineer next-generation bionanomaterials. In this Review, we analyse the background physics of several clinically relevant imaging modalities and their associated sensitivity and specificity, provide an overview of the materials currently used for in vivo nanodiagnostics, and assess the progress made towards clinical translation. This work provides a framework for understanding both the impressive progress made thus far in the nanodiagnostics field as well as presenting challenges that must be overcome to obtain widespread clinical adoption.

Discriminating cellular heterogeneity using microwell-based RNA cytometry

Discriminating cellular heterogeneity is important for understanding cellular physiology. However, it is limited by the technical difficulties of single-cell measurements. Here we develop a two-stage system to determine cellular heterogeneity. In the first stage, we perform multiplex single-cell RNA cytometry in a microwell array containing over 60,000 reaction chambers. In the second stage, we use the RNA cytometry data to determine cellular heterogeneity by providing a heterogeneity likelihood score (HLS). Moreover, we use Monte-Carlo simulation and RNA cytometry data to calculate the minimum number of cells required for detecting heterogeneity. We apply this system to characterize the RNA distributions of ageing-related genes in a highly purified mouse haematopoietic stem cell population. We identify genes that reveal novel heterogeneity of these cells. We also show that changes in expression of genes such as Birc6 during ageing can be attributed to the shift of relative portions of cells in the high-expressing subgroup versus low-expressing subgroup.

Molecular profiling of single circulating tumor cells from lung cancer patients

Significance There exists an urgent need for minimally invasive molecular analysis tools for cancer assessment and management, particularly in advanced-stage lung cancer, when tissue procurement is challenging and gene mutation profiling is crucial to identify molecularly targeted agents for treatment. High-throughput compartmentalization and multigene profiling of individual circulating tumor cells (CTCs) from whole-blood samples using modular gene panels may facilitate highly sensitive, yet minimally invasive characterization of lung cancer for therapy prediction and monitoring. We envision this nanoplatform as a compelling research tool to investigate the dynamics of cancer disease processes, as well as a viable clinical platform for minimally invasive yet comprehensive cancer assessment. Circulating tumor cells (CTCs) are established cancer biomarkers for the “liquid biopsy” of tumors. Molecular analysis of single CTCs, which recapitulate primary and metastatic tumor biology, remains challenging because current platforms have limited throughput, are expensive, and are not easily translatable to the clinic. Here, we report a massively parallel, multigene-profiling nanoplatform to compartmentalize and analyze hundreds of single CTCs. After high-efficiency magnetic collection of CTC from blood, a single-cell nanowell array performs CTC mutation profiling using modular gene panels. Using this approach, we demonstrated multigene expression profiling of individual CTCs from non–small-cell lung cancer (NSCLC) patients with remarkable sensitivity. Thus, we report a high-throughput, multiplexed strategy for single-cell mutation profiling of individual lung cancer CTCs toward minimally invasive cancer therapy prediction and disease monitoring.

Toward Integrated Molecular Diagnostic System (

Integrated molecular diagnostic systems (iMDx), which are automated, sensitive, specific, user-friendly, robust, rapid, easy-to-use, and portable, can revolutionize future medicine. This review will first focus on the components of sample extraction, preservation, and filtration necessary for all point-of-care devices to include for practical use. Subsequently, we will look for low-powered and precise methods for both sample amplification and signal transduction, going in-depth to the details behind their principles. The final field of total device integration and its application to the clinical field will also be addressed to discuss the practicality for future patient care. We envision that microfluidic systems hold the potential to breakthrough the number of problems brought into the field of medical diagnosis today.

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Biomolecular transport in nanofluidic confinement offers various means to investigate the behavior of biomolecules in their native aqueous environments, and to develop tools for diverse single-molecule manipulations. Recently, a number of simple nanofluidic fabrication techniques has been demonstrated that utilize electrospun nanofibers as a backbone structure. These techniques are limited by the arbitrary dimension of the resulting nanochannels due to the random nature of electrospinning. Here, a new method for fabricating nanofluidic systems from size-reduced electrospun nanofibers is reported and demonstrated. As it is demonstrated, this method uses the scanned electrospinning technique for generation of oriented sacrificial nanofibers and exposes these nanofibers to harsh, but isotropic etching/heating environments to reduce their cross-sectional dimension. The creation of various nanofluidic systems as small as 20 nm is demonstrated, and practical examples of single biomolecular handling, such as DNA elongation in nanochannels and fluorescence correlation spectroscopic analysis of biomolecules passing through nanochannels, are provided.

MDx): Principles and Applications

We present the integration of a cyclo olefin copolymer microfluidic chip with electrochemical pumps and an SU-8 tip for electrospray ionization mass spectrometry. The electrochemical pump, using electrolysis as an internal pressure source, was fabricated directly on the surface of the polymer chip. The triangular SU-8 emitter tip was fabricated on a glass wafer using standard photolithography. After release from the glass wafer, this tip was aligned to the microchannel and bonded between two polymer plates. The electrochemical pump and the electrospray tip were tested with electrospray ionization mass spectrometry. Mass spectrometry confirmed the stability of the electrochemical pump and the electrospray tip.

Rapid Prototyping of Nanofluidic Systems Using Size-Reduced Electrospun Nanofibers for Biomolecular Analysis

Resonant nanoelectromechanical systems have been demonstrated as sensitive mass detectors with subattogram and even single molecule sensitivity [Ilic et al., Nano Lett. 5, 925 (2005); Ilic et al., J. Appl. Phys. 95, 3694 (2004)]. Measurements of sub-ng/ml protein concentrations and DNA hybridization using deflection based microelectromechanical system (MEMS) devices have also been shown [Wu et al., Nat. Biotechnol. 19, 856 (2001); Fritz et al., Science 288, 316 (2000)]. Sample delivery is generally difficult in such cases requiring the entire device chip to be submersed into an analyte containing mixture. Additionally, in the case of MEMS resonators, high vacuum is required to remove viscous damping to improve sensitivity. In this work, the authors present a method where arrays of nanoelectromechanical devices are encapsulated in individually accessible, parallel microfluidic channels. The microchannels were used for delivery of liquids and nitrogen (for drying). The channels were pumped down to pressures...

On-chip coupling of electrochemical pumps and an SU-8 tip for electrospray ionization mass spectrometry

We present two novel methods to create microfluidic encapsulated nanoelectromechanical (NEMS) resonator arrays. NEMS arrays were encapsulated in individually accessible parallel microfluidic channels defined in glass. The channels could be filled with fluid as well as evacuated to sub-millitorr pressures for optical measurement of resonant device motion. Each 200 nm thick silicon nitride NEMS resonator was 3 mum wide with lengths ranging from 5 to 10 mum. The demonstrated resonant frequencies ranged from 3 to 10 MHz with quality factors of up to 3,500 (under vacuum). These devices showed no stiction after repeated wetting and drying while in the channels. The presented work demonstrates an important step towards a chip level total analytical system using NEMS devices in applications such as mass based biosensors