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Barrett K Duan, Peter E Cavanagh, Xiang Li, David R Walt
This report describes a novel method for isolating and detecting individual enzyme molecules using polymer arrays of picoliter microwells. A fluidic flow-cell device containing an array of microwells is fabricated in cyclic olefin polymer (COP). The use of COP microwell arrays simplifies experiments by eliminating extensive device preparation and surface functionalization that are common in other microwell array formats. Using a simple and robust loading method to introduce the reaction solution, individual enzyme molecules are trapped in picoliter microwells and subsequently isolated and sealed by fluorinated oil...
February 9, 2018: Analytical Chemistry
Jian-Zhang Pan, Pan Fang, Xiao-Xia Fang, Ting-Ting Hu, Jin Fang, Qun Fang
In this work, we developed a miniaturized palmtop high-speed capillary electrophoresis (CE) system integrating whole modules, including picoliter-scale sample injection, short capillary-based fast CE, high-voltage power supply, orthogonal laser induced fluorescence (LIF) detection, battery, system control, on-line data acquisition, processing, storage, and display modules. A strategy of minimalist miniaturization combining minimal system design and low-cost system construction was adopted to achieve the instrument miniaturization with extremely low cost, which is differing from the current microfabrication strategy used in most reported miniaturized CE systems...
January 29, 2018: Scientific Reports
Armend G Håti, Tomasz R Szymborski, Mathias Steinacher, Esther Amstad
Drops are often used as picoliter-sized reaction vessels, for example for high-throughput screening assays, or as templates to produce particles of controlled sizes and compositions. Many of these applications require close control over the size of drops, which can be achieved if they are produced with microfluidics. However, this tight size control comes at the expense of the throughput that is too low for many materials science and almost all industrial applications. To overcome this limitation, different parallelized microfluidic devices have been reported...
January 23, 2018: Lab on a Chip
A Ganguli, A Ornob, N Spegazzini, Y Liu, G Damhorst, T Ghonge, B Thornton, C J Konopka, W Dobrucki, S E Clare, R Bhargava, A M Smith, F Kosari, R Bashir
Here, we present a technique that performs on-chip picoliter real-time reverse transcriptase loop mediated isothermal amplification (RT-LAMP) reactions on a histological tissue section without any analyte purification while preserving the native spatial location of the nucleic acid molecules. We demonstrate this method by amplifying TOP2A messenger RNA (mRNA) in a prostate cancer xenograft with 100 µm spatial resolution and by visualizing the variation in threshold time of amplification across the tissue...
January 15, 2018: Nature Communications
Jinyu Chen, Zhaofeng Luo, Lin Li, Jinlong He, Luoquan Li, Jianwei Zhu, Ping Wu, Liqun He
The droplet digital polymerase chain reaction (ddPCR) is becoming more and more popular in diagnostic applications in academia and industry. In commercially available ddPCR systems, after they have been made by a generator, the droplets have to be transferred manually to modules for amplification and detection. In practice, some of the droplets (∼10%) are lost during manual transfer, leading to underestimation of the targets. In addition, the droplets are also at risk of cross-contamination during transfer...
January 5, 2018: Lab on a Chip
Lei Wu, Zhichao Dong, Fengyu Li, Yanlin Song
Manipulation of arrayed tiny droplets is important in liquid dispersion, liquid transportation, bioassays, nucleation, integrated electronics, and various lab experiments that require delivering precise and minute volumes of droplets. Liquid dispensed from a small orifice or split from surface patterns are typical methods, but the acquired droplet diameters are similar to that of the nozzle and pattern. Here we demonstrate that tiny droplets with dimensions much smaller than the pattern can be arrayed advantageously through designing a Laplace pressure pattern based on conical morphology and wetting heterogeneity...
January 16, 2018: Langmuir: the ACS Journal of Surfaces and Colloids
Richard W Rambach, Preetika Biswas, Ashutosh Yadav, Piotr Garstecki, Thomas Franke
The selective manipulation and incubation of individual picoliter drops in high-throughput droplet based microfluidic devices still remains challenging. We used a surface acoustic wave (SAW) to induce a bubble in a 3D designed multi-trap polydimethylsiloxane (PDMS) device to manipulate multiple droplets and demonstrate the selection, incubation and on-demand release of aqueous droplets from a continuous oil flow. By controlling the position of the acoustic actuation, individual droplets are addressed and selectively released from a droplet stream of 460 drops per s...
December 13, 2017: Analyst
Thomas Meier, Nan Wang, Dario Mager, Jan G Korvink, Sylvain Petitgirard, Leonid Dubrovinsky
A new pathway to nuclear magnetic resonance (NMR) spectroscopy for picoliter-sized samples (including those kept in harsh and extreme environments, particularly in diamond anvil cells) is introduced, using inductively coupled broadband passive electromagnetic lenses, to locally amplify the magnetic field at the isolated sample, leading to an increase in sensitivity. The lenses are adopted for the geometrical restrictions imposed by a toroidal diamond indenter cell and yield signal-to-noise ratios at pressures as high as 72 GPa at initial sample volumes of only 230 pl...
December 2017: Science Advances
Lucas R Parent, Evangelos Bakalis, Maria Proetto, Yiwen Li, Chiwoo Park, Francesco Zerbetto, Nathan C Gianneschi
Revolutions in science and engineering frequently result from the development, and wide adoption, of a new, powerful characterization or imaging technique. Beginning with the first glass lenses and telescopes in astronomy, to the development of visual-light microscopy, staining techniques, confocal microscopy, and fluorescence super-resolution microscopy in biology, and most recently aberration-corrected, cryogenic, and ultrafast (4D) electron microscopy, X-ray microscopy, and scanning probe microscopy in nanoscience...
December 11, 2017: Accounts of Chemical Research
Jessica Garcia, Eric Dusserre, Valérie Cheynet, Pierre Paul Bringuier, Karen Brengle-Pesce, Anne-Sophie Wozny, Claire Rodriguez-Lafrasse, Gilles Freyer, Marie Brevet, Léa Payen, Sébastien Couraud
Non invasive somatic detection assays are suitable for repetitive tumor characterization or for detecting the appearance of somatic resistance during lung cancer. Molecular diagnosis based on circulating free DNA (cfDNA) offers the opportunity to track the genomic evolution of the tumor, and was chosen to assess the molecular profile of several EGFR alterations, including deletions in exon 19 (delEX19), the L858R substitution on exon 21 and the EGFR resistance mutation T790M on exon 20. Our study aimed at determining optimal pre-analytical conditions and EGFR mutation detection assays for analyzing cfDNA using the picoliter-droplet digital polymerase chain reaction (ddPCR) assay...
October 20, 2017: Oncotarget
Gopakumar Kamalakshakurup, Abraham P Lee
Single cell analysis has emerged as a paradigm shift in cell biology to understand the heterogeneity of individual cells in a clone for pathological interrogation. Microfluidic droplet technology is a compelling platform to perform single cell analysis by encapsulating single cells inside picoliter-nanoliter (pL-nL) volume droplets. However, one of the primary challenges for droplet based single cell assays is single cell encapsulation in droplets, currently achieved either randomly, dictated by Poisson statistics, or by hydrodynamic techniques...
November 15, 2017: Lab on a Chip
Tianjin Yang, Stavros Stavrakis, Andrew deMello
Droplet-based microfluidic systems that incorporate flowing streams of pL-volume droplets surrounded by a continuous and immiscible carrier phase have attracted significant recent attention due to their utility in complex chemical and biological experimentation. Analysis of pL droplets, generated at kHz frequencies and moving at high linear velocities, is almost exclusively achieved using fluorescence-based detection schemes. To extend the applicability of such optical detection schemes, we herein report the development of a simple and cost-effective optofluidic platform, integrating liquid-core PDMS waveguides, that allows the accurate measurement of absorbance within individual pL-volume droplets moving within segmented flows...
November 17, 2017: Analytical Chemistry
Qian Yu, Fei Huang, Meilin Zhang, Haiying Ji, Shenchao Wu, Ying Zhao, Chunyan Zhang, Jiong Wu, Beili Wang, Baisheng Pan, Xin Zhang, Wei Guo
To explore the possible diagnostic value of liquid biopsy, two multiplex panels using picoliter-droplet digital polymerase chain reaction (ddPCR) were established to quantitatively assess the epidermal growth factor receptor (EGFR) mutations in cell‑free DNA (cfDNA) extracted from the plasma of advanced non‑small cell lung cancer (NSCLC) patients. Plasma samples derived from 22 patients with stage IIIB/IV NSCLC harboring EGFR mutations in matched tumor tissues confirmed by amplification refractory mutation system (ARMS) analysis were subjected to two multiplex ddPCR panels to assess the abundance of tyrosine kinase inhibitor (TKI) ‑sensitive (19DEL, L858R) and TKI‑resistant (T790 M) mutations...
August 2017: Molecular Medicine Reports
Meng Qi, Marina C Philip, Ning Yang, Jonathan V Sweedler
Metabolomics, the characterization of metabolites and their changes within biological systems, has seen great technological and methodological progress over the past decade. Most metabolomic experiments involve the characterization of the small-molecule content of fluids or tissue homogenates. While these microliter and larger volume metabolomic measurements can characterize hundreds to thousands of compounds, the coverage of molecular content decreases as sample sizes are reduced to the nanoliter and even to the picoliter volume range...
October 19, 2017: ACS Chemical Neuroscience
Manuela Pfammatter, Maria Andreasen, Georg Meisl, Christopher G Taylor, Jozef Adamcik, Sreenath Bolisetty, Antoni Sánchez-Ferrer, David Klenerman, Christopher M Dobson, Raffaele Mezzenga, Tuomas P J Knowles, Adriano Aguzzi, Simone Hornemann
The self-replicating properties of proteins into amyloid fibrils is a common phenomenon and underlies a variety of neurodegenerative diseases. Because propagation-active fibrils are chemically indistinguishable from innocuous aggregates and monomeric precursors, their detection requires measurements of their replicative capacity. Here we present a digital amyloid quantitative assay (d-AQuA) with insulin as model protein for the absolute quantification of single replicative units, propagons. D-AQuA is a microfluidics-based technology that performs miniaturized simultaneous propagon-induced amplification chain reactions within hundreds to thousands of picoliter-sized droplets...
November 1, 2017: Analytical Chemistry
Richard M Maceiczyk, David Hess, Flora W Y Chiu, Stavros Stavrakis, Andrew J deMello
Despite the growing importance of droplet-based microfluidics in high-throughput experimentation, few current methods allow the sensitive measurement of absorbance within rapidly moving droplets. To address this significant limitation, we herein present the application of differential detection photothermal interferometry (DDPI) for single-point absorbance quantification in pL- and fL-volume droplets. To assess the efficacy of our approach, we initially measure absorbance in 100 pL droplets at frequencies in excess of 1 kHz and determine a detection limit of 1...
October 25, 2017: Lab on a Chip
T S Kaminski, P Garstecki
Droplet microfluidics is a relatively new and rapidly evolving field of science focused on studying the hydrodynamics and properties of biphasic flows at the microscale, and on the development of systems for practical applications in chemistry, biology and materials science. Microdroplets present several unique characteristics of interest to a broader research community. The main distinguishing features include (i) large numbers of isolated compartments of tiny volumes that are ideal for single cell or single molecule assays, (ii) rapid mixing and negligible thermal inertia that all provide excellent control over reaction conditions, and (iii) the presence of two immiscible liquids and the interface between them that enables new or exotic processes (the synthesis of new functional materials and structures that are otherwise difficult to obtain, studies of the functions and properties of lipid and polymer membranes and execution of reactions at liquid-liquid interfaces)...
October 16, 2017: Chemical Society Reviews
Mohsen Karbaschi, Payam Shahi, Adam R Abate
Droplet microfluidics can form and process millions of picoliter droplets with speed and ease, allowing the execution of huge numbers of biological reactions for high-throughput studies. However, at the conclusion of most experiments, the emulsions must be broken to recover and analyze their contents. This is usually achieved with demulsifiers, like perfluorooctanol and chloroform, which can interfere with downstream reactions and harm cells. Here, we describe a simple approach to rapidly and efficiently break microfluidic emulsions, which requires no chemicals...
July 2017: Biomicrofluidics
P Kehayias, A Jarmola, N Mosavian, I Fescenko, F M Benito, A Laraoui, J Smits, L Bougas, D Budker, A Neumann, S R J Brueck, V M Acosta
Sensors using nitrogen-vacancy centers in diamond are a promising tool for small-volume nuclear magnetic resonance (NMR) spectroscopy, but the limited sensitivity remains a challenge. Here we show nearly two orders of magnitude improvement in concentration sensitivity over previous nitrogen-vacancy and picoliter NMR studies. We demonstrate NMR spectroscopy of picoliter-volume solutions using a nanostructured diamond chip with dense, high-aspect-ratio nanogratings, enhancing the surface area by 15 times. The nanograting sidewalls are doped with nitrogen-vacancies located a few nanometers from the diamond surface to detect the NMR spectrum of roughly 1 pl of fluid lying within adjacent nanograting grooves...
August 4, 2017: Nature Communications
Russell H Cole, Shi-Yang Tang, Christian A Siltanen, Payam Shahi, Jesse Q Zhang, Sean Poust, Zev J Gartner, Adam R Abate
Although the elementary unit of biology is the cell, high-throughput methods for the microscale manipulation of cells and reagents are limited. The existing options either are slow, lack single-cell specificity, or use fluid volumes out of scale with those of cells. Here we present printed droplet microfluidics, a technology to dispense picoliter droplets and cells with deterministic control. The core technology is a fluorescence-activated droplet sorter coupled to a specialized substrate that together act as a picoliter droplet and single-cell printer, enabling high-throughput generation of intricate arrays of droplets, cells, and microparticles...
August 15, 2017: Proceedings of the National Academy of Sciences of the United States of America
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