189 research outputs found
Local detection of enzymatic ion generation with polycrystalline silicon interdigitated electrodes and its application to biosensing
Chips containing polycrystalline silicon interdigitated electrodes are modified with the enzyme urease. The sensors are able to detect changes in the resitivity of the solution near their surface, where the enzymatic reaction generates charged species. The electrodes are also grafted with an antigen and queried with different amounts of urease labeled antibody. The response of the modified electrodes is proportional to the amount of enzyme attached to the surface by the biorecognition event, thus validating the assay for biosensing applications
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Live single cell functional phenotyping in droplet nano-liter reactors
While single cell heterogeneity is present in all biological systems, most studies cannot address it due to technical limitations. Here we describe a nano-liter droplet microfluidic-based approach for stimulation and monitoring of surfaceand secreted markers of live single immune dendritic cells (DCs) as well as monitoring the live T cell/DC interaction. This nano-liter in vivo simulating microenvironment allows delivering various stimuli reagents to each cell and appropriate gas exchanges which are necessary to ensure functionality and viability of encapsulated cells. Labeling bioassay and microsphere sensors were integrated into nano-liter reaction volume of the droplet to monitor live single cell surface markers and secretion analysis in the time-dependent fashion. Thus live cell stimulation, secretion and surface monitoring can be obtained simultaneously in distinct microenvironment, which previously was possible using complicated and multi-step in vitro and in vivo live-cell microscopy, together with immunological studies of the outcome secretion of cellular function
Longitudinal multiparameter assay of lymphocyte interactions from onset by microfluidic cell pairing and culture
Resolving how the early signaling events initiated by cell–cell interactions are transduced into diverse functional outcomes necessitates correlated measurements at various stages. Typical approaches that rely on bulk cocultures and population-wide correlations, however, only reveal these relationships broadly at the population level, not within each individual cell. Here, we present a microfluidics-based cell–cell interaction assay that enables longitudinal investigation of lymphocyte interactions at the single-cell level through microfluidic cell pairing, on-chip culture, and multiparameter assays, and allows recovery of desired cell pairs by micromanipulation for off-chip culture and analyses. Well-defined initiation of interactions enables probing cellular responses from the very onset, permitting single-cell correlation analyses between early signaling dynamics and later-stage functional outcomes within same cells. We demonstrate the utility of this microfluidic assay with natural killer cells interacting with tumor cells, and our findings suggest a possible role for the strength of early calcium signaling in selective coordination of subsequent cytotoxicity and IFN-gamma production. Collectively, our experiments demonstrate that this new approach is well-suited for resolving the relationships between complex immune responses within each individual cell.Singapore-MIT AllianceAmerican Association for Cancer Research. Pancreatic Cancer Action NetworkMassachusetts Institute of Technology. Department of Electrical Engineering and Computer Science (Frank Quick Faculty Research Innovation Fellowship
Advances in Microfluidics and Lab-on-a-Chip Technologies
Advances in molecular biology are enabling rapid and efficient analyses for
effective intervention in domains such as biology research, infectious disease
management, food safety, and biodefense. The emergence of microfluidics and
nanotechnologies has enabled both new capabilities and instrument sizes
practical for point-of-care. It has also introduced new functionality, enhanced
sensitivity, and reduced the time and cost involved in conventional molecular
diagnostic techniques. This chapter reviews the application of microfluidics
for molecular diagnostics methods such as nucleic acid amplification,
next-generation sequencing, high resolution melting analysis, cytogenetics,
protein detection and analysis, and cell sorting. We also review microfluidic
sample preparation platforms applied to molecular diagnostics and targeted to
sample-in, answer-out capabilities
Nanowell-Based Immunoassays for Measuring Single-Cell Secretion: Characterization of Transport and Surface Binding
Arrays of subnanoliter wells (nanowells) provide a useful system to isolate single cells and analyze their secreted proteins. Two general approaches have emerged: one that uses open arrays and local capture of secreted proteins, and a second (called microengraving) that relies on closed arrays to capture secreted proteins on a solid substrate, which is subsequently removed from the array. However, the design and operating parameters for efficient capture from these two approaches to analyze single-cell secretion have not been extensively considered. Using numerical simulations, we analyzed the operational envelope for both open and closed formats, as a function of the spatial distribution of capture ligands, their affinities for the protein, and the rates of single-cell secretion. Based on these analyses, we present a modified approach to capture secreted proteins in-well for highly active secreting cells. This simple method for in-well detection should facilitate rapid identification of cell lines with high specific productivities.National Institutes of Health (U.S.)/National Institute of Allergy and Infectious Diseases (U.S.) (5P01AI045757)National Cancer Institute (U.S.) (Koch Institute Support (Core) Grant P30-CA14051
An All-Photonic Molecule-Based D Flip-Flop
The photochromic fluorescence switching of a fulgimide derivative was used to implement the first molecule-based D (delay) flip-flop device, which works based on the principles of sequential logic. The device operates exclusively with photonic signals and can be conveniently switched in repeated cycles. \ua9 2011 American Chemical Society
OFF-ON-OFF Fluorescence Switch with T-Latch Function
A novel molecular system with characteristics of an OFF-ON-OFF fluorescence switch was designed to integrate the function of a T-latch. In detail, a receptor(1)-fluorophore-receptor(2) architecture was adopted to achieve fluorescence switching upon addition of protons
Porous Bead-Based Diagnostic Platforms: Bridging the Gaps in Healthcare
Advances in lab-on-a-chip systems have strong potential for multiplexed
detection of a wide range of analytes with reduced sample and reagent volume; lower costs
and shorter analysis times. The completion of high-fidelity multiplexed and multiclass
assays remains a challenge for the medical microdevice field; as it struggles to achieve and
expand upon at the point-of-care the quality of results that are achieved now routinely in
remote laboratory settings. This review article serves to explore for the first time the key
intersection of multiplexed bead-based detection systems with integrated microfluidic
structures alongside porous capture elements together with biomarker validation studies.
These strategically important elements are evaluated here in the context of platform
generation as suitable for near-patient testing. Essential issues related to the scalability of
these modular sensor ensembles are explored as are attempts to move such multiplexed and
multiclass platforms into large-scale clinical trials. Recent efforts in these bead sensors
have shown advantages over planar microarrays in terms of their capacity to generate
multiplexed test results with shorter analysis times. Through high surface-to-volume ratios
and encoding capabilities; porous bead-based ensembles; when combined with
microfluidic elements; allow for high-throughput testing for enzymatic assays; general
chemistries; protein; antibody and oligonucleotide applications
All-Photonic Multifunctional Molecular Logic Device
Photochromes are photoswitchable, bistable chromophores which, like transistors, can implement binary logic operations. When several photochromes are combined in one molecule, interactions between them such as energy and electron transfer allow design of simple Boolean logic gates and more complex logic devices with all-photonic inputs and outputs. Selective isomerization of individual photochromes can be achieved using light of different wavelengths, and logic outputs can employ absorption and emission properties at different wavelengths, thus allowing a single molecular species to perform several different functions, even simultaneously. Here, we report a molecule consisting of three linked photochromes that can be configured as AND, XOR, INH, half-adder, half-subtractor, multiplexer, demultiplexer, encoder, decoder, keypad lock, and logically reversible transfer gate logic devices, all with a common initial state. The system demonstrates the advantages of light-responsive molecules as multifunctional, reconfigurable nanoscale logic devices that represent an approach to true molecular information processing units
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