Single-cell whole-transcriptome analysis is a powerful tool for quantifying gene expression heterogeneity in populations of cells. Many techniques have, thus, been recently developed to perform transcriptome sequencing (RNA-Seq) on individual cells. To probe subtle biological variation between samples with limiting amounts of RNA, more precise and sensitive methods are still required. We adapted a previously developed strategy for single-cell RNA-Seq that has shown promise for superior sensitivity and implemented the chemistry in a microfluidic platform for single-cell wholetranscriptome analysis. In this approach, single cells are captured and lysed in a microfluidic device, where mRNAs with poly(A) tails are reverse-transcribed into cDNA. Double-stranded cDNA is then collected and sequenced using a next generation sequencing platform. We prepared 94 libraries consisting of single mouse embryonic cells and technical replicates of extracted RNA and thoroughly characterized the performance of this technology. Microfluidic implementation increased mRNA detection sensitivity as well as improved measurement precision compared with tube-based protocols. With 0.2 M reads per cell, we were able to reconstruct a majority of the bulk transcriptome with 10 single cells. We also quantified variation between and within different types of mouse embryonic cells and found that enhanced measurement precision, detection sensitivity, and experimental throughput aided the distinction between biological variability and technical noise. With this work, we validated the advantages of an early approach to single-cell RNA-Seq and showed that the benefits of combining microfluidic technology with high-throughput sequencing will be valuable for large-scale efforts in single-cell transcriptome analysis.genomics | lab on chip | embryonic stem cell
This study reports a microfluidic chip-based wearable colorimetric sensor for detecting sweat glucose. The device consisted of five microfluidic channels branching out from the center and connected to the detection microchambers. The microchannels could route the sweat excreted from the epidermis to the microchambers, and each of them was integrated with a check valve to avoid the risk of the backflow of the chemical reagents from the microchamber. The microchambers contained the pre-embedded glucose oxidase (GOD)–peroxidase–o-dianisidine reagents for sensing the glucose in sweat. It was found that the color change caused by the enzymatic oxidation of o-dianisidine could show a more sensitive response to the glucose than that of the conventional GOD–peroxidase–KI system. This sensor could perform five parallel detections at one time. The obtained linear range for sweat glucose was 0.1–0.5 mM with a limit of detection of 0.03 mM. The sensor was also used to detect the glucose in sweat samples from a group of subjects engaged in both fasting and postprandial trials. The results showed that our wearable colorimetric sensor can reveal the subtle differences existing in the sweat glucose concentration after the fasting and the oral glucose uptake.
Carbonation of natural brucite in H2O and diluted HCl is investigated at room temperature and moderate pCO2 to explore the products' mineralogy and reaction kinetics. Results show nesquehonite is by far the dominant carbonate species formed, despite its poorer thermodynamic stability relative to magnesite and possibly hydromagnesite. Time-dependent measurements reveal carbonate formation within 30 min, regardless of the original acidity of the slurry. However, while the fraction of reacted brucite in H2O increases gradually over time and approaches unity ( approximately 98%) at 2.5 h, it rises rapidly in HCl within the first hour and levels off thereafter, leaving a significant amount of brucite unreacted. Such behavior suggests that the initial quantity of Mg2+ affects the reaction kinetics. Fitting a pseudo first-order rate law to the data yields a higher rate constant for the HCl experiments. These observations may imply that the carbonation does not proceed through heterogeneous reaction between gaseous CO2 and solid brucite. Solution chemistry analysis indicates that most CO2 stays in aqueous phase in both media; however, the concentration of HCO3(-) becomes high in H2O after about 2 h, agreeing with the observed inferior carbonation extent in HCl.
The interaction between tumor and endothelial cells is crucial to cancer metastasis and angiogenesis. We developed a novel microfluidic device to assess the cell-cell interaction quantitatively at the single cell resolution. This integrated chip offers 16 coculture experiments in parallel with controllable microenvironments to study interactions between cells dynamically. We applied this approach to model the tumor invasion using Hela cells and human umbilical vein endothelial cells (HUVECs) and monitored the migration of both. We observed the retreatment of HUVECs upon the approach of Hela cells during coculture, indicating that the interaction between two cells was mediated by soluble factors. This interaction was further analyzed through quantitatively processing the phase-contrast microscopic time-lapse images of each individual coculture chamber. We also confirmed this paracrine effect by varying the frequency of medium change. This microfluidic technique is highly controllable, contamination free, fully automatic, and inexpensive. This approach not only offers a unique way to quantitatively study the interaction between cells but also provides accurate spatial-temporal tunability of microenvironments for cell coculture. We believe this method, intrinsically high-throughput and quantitative, will greatly facilitate the study of cell-cell interactions and communications.
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