A Multiwell-Based Detection Platform with Integrated PDMS Concentrators for Rapid Multiplexed Enzymatic Assays

We report an integrated system for accelerating assays with concentrators in a standard 12-well plate (ISAAC-12) and demonstrate its versatility for rapid detection of matrix metalloproteinase (MMP)-9 expression in the cell culture supernatant of breast cancer cell line MDA-MB-231 by accelerating the enzymatic reaction and end-point signal intensity via electrokinetic preconcentration. Using direct printing of a conductive ion-permselective polymer on a polydimethylsiloxane (PDMS) channel, the new microfluidic concentrator chip can be built without modifying the underlying substrate. Through this decoupling fabrication strategy, our microfluidic concentrator chip can easily be integrated with a standard multiwell plate, the de facto laboratory standard platform for high-throughput assays, simply by reversible bonding on the bottom of each well. It increases the reaction rate of enzymatic assays by concentrating the enzyme and the reaction product inside each well simultaneously for rapid multiplexed detection.publishersversionpublishe

Capillary-valve-based fabrication of ion-selective membrane junction for electrokinetic sample preconcentration in PDMS chip

In this paper, we report a novel method for fabricating ion-selective membranes in poly(dimethylsiloxane) (PDMS)/glass-based microfluidic preconcentrators. Based on the concept of capillary valves, this fabrication method involves filling a lithographically patterned junction between two microchannels with an ion-selective material such as Nafion resin; subsequent curing results in a high aspect-ratio membrane for use in electrokinetic sample preconcentration. To demonstrate the concentration performance of this high-aspect-ratio, ion-selective membrane, we integrated the preconcentrator with a surface-based immunoassay for R-Phycoerythrin (RPE). Using a 1× PBS buffer system, the preconcentrator-enhanced immunoassay showed an approximately 100× improvement in sensitivity within 30 min. This is the first time that an electrokinetic microfluidic preconcentrator based on ion concentration polarization (ICP) has been used in high ionic strength buffer solutions to enhance the sensitivity of a surface-based immunoassay.National Institutes of Health (U.S.) (CA119402)National Institutes of Health (U.S.) (EB005743

Nanofluidic Concentration Device for Biomolecules Utilizing Ion Concentration Polarization: Theory, Fabrication, and Applications

This article was published as part of the From microfluidic application to nanofluidic phenomena issueRecently, a new type of electrokinetic concentration devices has been developed in a microfluidic chip format, which allows efficient trapping and concentration of biomolecules by utilizing ion concentration polarization near nanofluidic structures. These devices have drawn much attention not only due to their potential application in biomolecule sensing, but also due to the rich scientific content related to ion concentration polarization, the underlying physical phenomenon for the operation of these electrokinetic concentration devices. This tutorial review provides an introduction to the scientific and engineering advances achieved, in-depth discussion about several interesting applications of these unique concentration devices, and their current limitations and challenges.National Science Foundation (U.S.) (CBET-0347348 & 0854026)National Institutes of Health (U.S.) (Grant EB005743)National Institutes of Health (U.S.) (Grant CA119402)National Institutes of Health (U.S.) (Grant P50-GM68762

Graphene Photonics and Optoelectronics

The richness of optical and electronic properties of graphene attracts enormous interest. Graphene has high mobility and optical transparency, in addition to flexibility, robustness and environmental stability. So far, the main focus has been on fundamental physics and electronic devices. However, we believe its true potential to be in photonics and optoelectronics, where the combination of its unique optical and electronic properties can be fully exploited, even in the absence of a bandgap, and the linear dispersion of the Dirac electrons enables ultra-wide-band tunability. The rise of graphene in photonics and optoelectronics is shown by several recent results, ranging from solar cells and light emitting devices, to touch screens, photodetectors and ultrafast lasers. Here we review the state of the art in this emerging field.Comment: Review Nature Photonics, in pres

Electrochemical activation and inhibition of neuromuscular systems through modulation of ion concentrations with ion-selective membranes

Conventional functional electrical stimulation aims to restore functional motor activity of patients with disabilities resulting from spinal cord injury or neurological disorders. However, intervention with functional electrical stimulation in neurological diseases lacks an effective implantable method that suppresses unwanted nerve signals. We have developed an electrochemical method to activate and inhibit a nerve by electrically modulating ion concentrations in situ along the nerve. Using ion-selective membranes to achieve different excitability states of the nerve, we observe either a reduction of the electrical threshold for stimulation by up to approximately 40%, or voluntary, reversible inhibition of nerve signal propagation. This low-threshold electrochemical stimulation method is applicable in current implantable neuroprosthetic devices, whereas the on-demand nerve-blocking mechanism could offer effective clinical intervention in disease states caused by uncontrolled nerve activation, such as epilepsy and chronic pain syndromes.Massachusetts Institute of Technology. Faculty Discretionary Research FundNational Institutes of Health (U.S.) (Award UL1 RR 025758)Harvard Catalyst (Grant

Chicken Pleiotrophin: Regulation of Tissue Specific Expression by Estrogen in the Oviduct and Distinct Expression Pattern in the Ovarian Carcinomas

Pleiotrophin (PTN) is a developmentally-regulated growth factor which is widely distributed in various tissues and also detected in many kinds of carcinomas. However, little is known about the PTN gene in chickens. In the present study, we found chicken PTN to be highly conserved with respect to mammalian PTN genes (91–92.6%) and its mRNA was most abundant in brain, heart and oviduct. This study focused on the PTN gene in the oviduct where it was detected in the glandular (GE) and luminal (LE) epithelial cells. Treatment of young chicks with diethylstilbesterol induced PTN mRNA and protein in GE and LE, but not in other cell types of the oviduct. Further, several microRNAs, specifically miR-499 and miR-1709 were discovered to influence PTN expression via its 3′-UTR which suggests that post-transcriptional regulation influences PTN expression in chickens. We also compared expression patterns and CpG methylation status of the PTN gene in normal and cancerous ovaries from chickens. Our results indicated that PTN is most abundant in the GE of adenocarcinoma of cancerous, but not normal ovaries of hens. Bisulfite sequencing revealed that 30- and 40% of −1311 and −1339 CpG sites are demethylated in ovarian cancer cells, respectively. Collectively, these results indicate that chicken PTN is a novel estrogen-induced gene expressed mainly in the oviductal epithelia implicating PTN regulation of oviduct development and egg formation, and also suggest that PTN is a biomarker for epithelial ovarian carcinoma that could be used for diagnosis and monitoring effects of therapies for the disease

Enhancing Protease Activity Assay in Droplet-Based Microfluidics Using a Biomolecule Concentrator

We introduce an integrated microfluidic device consisting of a biomolecule concentrator and a microdroplet generator, which enhances the limited sensitivity of low-abundance enzyme assays by concentrating biomolecules before encapsulating them into droplet microreactors. We used this platform to detect ultralow levels of matrix metalloproteinases (MMPs) from diluted cellular supernatant and showed that it significantly (~10-fold) reduced the time required to complete the assay and the sample volume used.National Institutes of Health (U.S.) (Grant GM68762)National Institutes of Health (U.S.) (Grant U54-CA112967)National Institutes of Health (U.S.) (Grant R01-EB010246)National Institutes of Health (U.S.) (Grant R01-GM081336)National Science Foundation (U.S.) (Graduate Fellowship)United States. Defense Advanced Research Projects Agency (Cipher Program

CRL4 antagonizes SCFFbxo7-mediated turnover of cereblon and BK channel to regulate learning and memory

Intellectual disability (ID), one of the most common human developmental disorders, can be caused by genetic mutations in Cullin 4B (Cul4B) and cereblon (CRBN). CRBN is a substrate receptor for the Cul4A/B-DDB1 ubiquitin ligase (CRL4) and can target voltage- and calcium-activated BK channel for ER retention. Here we report that ID-associated CRL4CRBNmutations abolish the interaction of the BK channel with CRL4, and redirect the BK channel to the SCFFbxo7ubiquitin ligase for proteasomal degradation. Glioma cell lines harbouring CRBN mutations record density-dependent decrease of BK currents, which can be restored by blocking Cullin ubiquitin ligase activity. Importantly, mice with neuron-specific deletion of DDB1 or CRBN express reduced BK protein levels in the brain, and exhibit similar impairment in learning and memory, a deficit that can be partially rescued by activating the BK channel. Our results reveal a competitive targeting of the BK channel by two ubiquitin ligases to achieve exquisite control of its stability, and support changes in neuronal excitability as a common pathogenic mechanism underlying CRL4CRBN–associated ID

Survey of childhood empyema in Asia: Implications for detecting the unmeasured burden of culture-negative bacterial disease

<p>Abstract</p> <p>Background</p> <p>Parapneumonic empyema continues to be a disease of significant morbidity and mortality among children despite recent advances in medical management. To date, only a limited number of studies have assessed the burden of empyema in Asia.</p> <p>Methods</p> <p>We surveyed medical records of four representative large pediatric hospitals in China, Korea, Taiwan and Vietnam using <it>ICD</it>-10 diagnostic codes to identify children <16 years of age hospitalized with empyema or pleural effusion from 1995 to 2005. We also accessed microbiology records of cultured empyema and pleural effusion specimens to describe the trends in the epidemiology and microbiology of empyema.</p> <p>Results</p> <p>During the study period, we identified 1,379 children diagnosed with empyema or pleural effusion (China, n = 461; Korea, n = 134; Taiwan, n = 119; Vietnam, n = 665). Diagnoses of pleural effusion (n = 1,074) were 3.5 times more common than of empyema (n = 305), although the relative proportions of empyema and pleural effusion noted in hospital records varied widely between the four sites, most likely because of marked differences in coding practices. Although pleural effusions were reported more often than empyema, children with empyema were more likely to have a cultured pathogen. In addition, we found that median age and gender distribution of children with these conditions were similar across the four countries. Among 1,379 empyema and pleural effusion specimens, 401 (29%) were culture positive. <it>Staphylococcus aureus </it>(n = 126) was the most common organism isolated, followed by <it>Streptococcus pneumoniae </it>(n = 83), <it>Pseudomonas aeruginosa </it>(n = 37) and <it>Klebsiella </it>(n = 35) and <it>Acinetobacter </it>species (n = 34).</p> <p>Conclusion</p> <p>The age and gender distribution of empyema and pleural effusion in children in these countries are similar to the US and Western Europe. <it>S. pneumoniae </it>was the second leading bacterial cause of empyema and pleural effusion among Asian children. The high proportion of culture-negative specimens among patients with pleural effusion or empyema suggests that culture may not be a sufficiently sensitive diagnostic method to determine etiology in the majority of cases. Future prospective studies in different countries would benefit from standardized case definitions and coding practices for empyema. In addition, more sensitive diagnostic methods would improve detection of pathogens and could result in better prevention, treatment and outcomes of this severe disease.</p