scholarly journals High-throughput, non-equilibrium studies of single biomolecules using glass-made nanofluidic devices

Lab on a Chip ◽  
2019 ◽  
Vol 19 (1) ◽  
pp. 79-86 ◽  
Author(s):  
Mattia Fontana ◽  
Carel Fijen ◽  
Serge G. Lemay ◽  
Klaus Mathwig ◽  
Johannes Hohlbein
Keyword(s):  
High Throughput ◽  
Single Molecule ◽  
Single Molecule Detection ◽  
Dynamic Heterogeneity ◽  
Equilibrium Studies ◽  
Complex Samples ◽  
Powerful Means ◽  
Nanofluidic Devices ◽  
Detection Schemes ◽  
Non Equilibrium

Single-molecule detection schemes offer powerful means to overcome static and dynamic heterogeneity inherent to complex samples.

scholarly journals Simple nanofluidic devices for high-throughput, non-equilibrium studies at the single-molecule level

2017 ◽  
Author(s):  
Carel Fijen ◽  
Mattia Fontana ◽  
Serge G. Lemay ◽  
Klaus Mathwig ◽  
Johannes Hohlbein
Keyword(s):  
High Throughput ◽  
Single Molecule ◽  
Single Particle Tracking ◽  
Biological Interactions ◽  
Dynamic Heterogeneity ◽  
High Throughput Analysis ◽  
Equilibrium Studies ◽  
Single Molecule Level ◽  
Nanofluidic Devices ◽  
And Performance

ABSTRACTSingle-molecule detection schemes offer powerful means to overcome static and dynamic heterogeneity inherent to complex samples. Probing chemical and biological interactions and reactions with high throughput and time resolution, however, remains challenging and often requires surface-immobilized entities. Here, utilizing camera-based fluorescence microscopy, we present glass-made nanofluidic devices in which fluorescently labelled molecules flow through nanochannels that confine their diffusional movement. The first design features an array of parallel nanochannels for high-throughput analysis of molecular species under equilibrium conditions allowing us to record 200.000 individual localization events in just 10 minutes. Using these localizations for single particle tracking, we were able to obtain accurate flow profiles including flow speeds and diffusion coefficients inside the channels.A second design featuring a T-shaped nanochannel enables precise mixing of two different species as well as the continuous observation of chemical reactions. We utilized the design to visualize enzymatically driven DNA synthesis in real time and at the single-molecule level. Based on our results, we are convinced that the versatility and performance of the nanofluidic devices will enable numerous applications in the life sciences.

Highly Sensitive Detection of Paraquat with Pillar[5]arene as an aptamer in α-Hemolysin Nanopore

2021 ◽  
Author(s):  
Xiaojia Jiang ◽  
Mingsong Zang ◽  
Fei Li ◽  
Chunxi Hou ◽  
Quan Luo ◽  
...  
Keyword(s):  
Real Time ◽  
High Throughput ◽  
Single Molecule ◽  
Single Molecule Detection ◽  
Sensitive Detection ◽  
High Throughput Analysis ◽  
Throughput Analysis ◽  
The Real ◽  
Highly Sensitive ◽  
Highly Sensitive Detection

Biological nanopore-based techniques have attracted more and more attention recently in the field of single-molecule detection, because they allow the real-time, sensitive, high-throughput analysis. Herein, we report an engineered biological...

Single-Molecule Detection Technologies in Miniaturized High-Throughput Screening: Fluorescence Intensity Distribution Analysis

2003 ◽  
Vol 8 (1) ◽  
pp. 19-33 ◽  
Author(s):  
Ulrich Haupts ◽  
Martin Rüdiger ◽  
Stephen Ashman ◽  
Sandra Turconi ◽  
Ryan Bingham ◽  
...  
Keyword(s):  
High Throughput ◽  
Single Molecule ◽  
Fluorescence Intensity ◽  
Intensity Distribution ◽  
High Throughput Screening ◽  
Biological Data ◽  
Single Molecule Detection ◽  
Distribution Analysis ◽  
Fluorescence Intensity Distribution Analysis ◽  
Detection Technologies

Single-molecule detection technologies are becoming a powerful readout format to support ultra-high-throughput screening. These methods are based on the analysis of fluorescence intensity fluctuations detected from a small confocal volume element. The fluctuating signal contains information about the mass and brightness of the different species in a mixture. The authors demonstrate a number of applications of fluorescence intensity distribution analysis (FIDA), which discriminates molecules by their specific brightness. Examples for assays based on brightness changes induced by quenching/dequenching of fluorescence, fluorescence energy transfer, and multiple-binding stoichiometry are given for important drug targets such as kinases and proteases. FIDA also provides a powerful method to extract correct biological data in the presence of compound fluorescence. ( Journal of Biomolecular Screening 2003:19-33)

scholarly journals Single Molecule Detection Technologies in Miniaturized High Throughput Screening: Fluorescence Correlation Spectroscopy

1999 ◽  
Vol 4 (6) ◽  
pp. 335-353 ◽  
Author(s):  
Keith J. Moore ◽  
Sandra Turconi ◽  
Stephen Ashman ◽  
Martin Ruediger ◽  
Ulrich Haupts ◽  
...  
Keyword(s):  
High Throughput ◽  
Single Molecule ◽  
Fluorescence Correlation Spectroscopy ◽  
High Throughput Screening ◽  
Equilibrium Constants ◽  
Correlation Spectroscopy ◽  
Single Molecule Detection ◽  
General Applicability ◽  
Detection Techniques ◽  
Fluorescence Correlation

Fluorescence assay technologies used for miniaturized high throughput screening are broadly divided into two classes. Macroscopic fluorescence techniques (encompassing conventional fluorescence intensity, anisotropy [also often referred to as fluorescence polarization] and energy transfer) monitor the assay volume- and time-averaged fluorescence output from the ensemble of emitting fluorophores. In contrast, single-molecule detection (SMD) techniques and related approaches, such as fluorescence correlation spectroscopy (FCS), stochastically sample the fluorescence properties of individual constituent molecules and only then average many such detection events to define the properties of the assay system as a whole. Analysis of single molecular events is accomplished using confocal optics with an illumination/detection volume of -1 fl (10-15 L) such that the signal is insensitive to miniaturization of HTS assays to 1 A1 or below. In this report we demonstrate the general applicability of one SMD technique (FCS) to assay configuration for target classes typically encountered in HTS and confirm the equivalence of the rate/equilibrium constants determined by FCS and by macroscopic techniques. Advantages and limitations of the current FCS technology, as applied here, and potential solutions, particularly involving alternative SMD detection techniques, are also discussed.

scholarly journals Spontaneous DNA Translocation Through van der Waals Heterostructure Nanopore for Single-Molecule Detection

2021 ◽  
Author(s):  
Yang Liu ◽  
Ye Deng ◽  
Yanmei Yang ◽  
Yuanyuan Qu ◽  
Chao Zhang ◽  
...  
Keyword(s):  
Solid State ◽  
High Precision ◽  
High Throughput ◽  
Single Molecule ◽  
Van Der Waals ◽  
Single Molecule Detection ◽  
Dna Translocation ◽  
New Paradigm ◽  
Van Der Waals Heterostructure ◽  
Long Read

Solid-state nanopore detection and sequencing of single-molecule offers a new paradigm because of several well-recognized features like long-read, high throughput, high precision and direct analyses. However, several key technical challenges...

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