Synthesis of new hydrophilic rhodamine based enzymatic substrates compatible with droplet-based microfluidic assays

Fenneteau, Johan; Chauvin, Dany; Griffiths, Andrew D.; Nizak, Clément; Cossy, J.

doi:10.1039/c7cc01506b

Cited by 24 publications

(20 citation statements)

References 35 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…One example is the synthesis of new hydrophilic substrates for the enzymatic reactions, which are characterized by minimized leakage from aqueous droplets to the oil. 50 In another collaborative project between our microfluidic research group and analytical chemists, we developed the system for monitoring the oxygen concentration and conditions for bacterial growth inside nanoliter trains of droplets. 39 The method took advantage of dedicated oxygen-sensitive indicator dyes embedded in nanoparticles.…”

Section: New Probes and Sensors For Optical Detectionmentioning

confidence: 99%

Controlled droplet microfluidic systems for multistep chemical and biological assays

2017

View full text Add to dashboard Cite

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). The most frequent application of droplet microfluidics relies on the generation of large numbers of compartments either for ultrahigh throughput screens or for the synthesis of functional materials composed of millions of droplets or particles. Droplet microfluidics has already evolved into a complex field. In this review we focus on 'controlled droplet microfluidics' - a portfolio of techniques that provide convenient platforms for multistep complex reaction protocols and that take advantage of automated and passive methods of fluid handling on a chip. 'Controlled droplet microfluidics' can be regarded as a group of methods capable of addressing and manipulating droplets in series. The functionality and complexity of controlled droplet microfluidic systems can be positioned between digital microfluidics (DMF) addressing each droplet individually using 2D arrays of electrodes and ultrahigh throughput droplet microfluidics focused on the generation of hundreds of thousands or even millions of picoliter droplets that cannot be individually addressed by their location on a chip.

show abstract

Section: New Probes and Sensors For Optical Detectionmentioning

confidence: 99%

Controlled droplet microfluidic systems for multistep chemical and biological assays

2017

View full text Add to dashboard Cite

show abstract

“…The droplet volume used in FADS ranges from several picoliter to hundreds of picoliter. To date, FADS has successfully applied for the screening of β-galactosidase, horseradish peroxidase, sulfatase, cellulase, aldolase, lipase, and others (Agresti et al 2010;Baret et al 2009;Fenneteau et al 2017;Kintses et al 2012;Obexer et al 2017;Ostafe et al 2014;Qiao et al 2018). The operation of FADS can be divided into the following steps (Baret et al 2009): 1) Droplet generation.…”

Section: High-throughput Single-cell Screening For Enzyme Discoverymentioning

confidence: 99%

“…The second is that the substrate must be effectively retained within the droplets with limited droplet cross-contamination (Skhiri et al 2012). Several methods have been developed to slow down the leakage and diffusion of small molecules, including fluorogenic substrates among droplets, such as modifying the fluorogenic substrates with higher hydrophilicity, improving the viscosity of the droplets by adding bovine serum albumin (BSA) or carboxymethyl cellulose (CMC) and using nanoparticles as a surfactant to avoid surfactant micelles transport of the substrate between droplets (Courtois et al 2009;Fenneteau et al 2017). Obexer et al (2017) developed a fully automated FADS device that integrated the modules of droplet generation, incubation and sorting on a single chip for directed evolution of a previously computationally designed enzyme into a highly active enzyme which requires screening of ~ 10 8 protein variants (Fig.…”

Section: High-throughput Single-cell Screening For Enzyme Discoverymentioning

confidence: 99%

One cell at a time: droplet-based microbial cultivation, screening and sequencing

et al. 2021

Mar Life Sci Technol

View full text Add to dashboard Cite

Microbes thrive and, in turn, influence the earth's environment, but most are poorly understood because of our limited capacity to reveal their natural diversity and function. Developing novel tools and effective strategies are critical to ease this dilemma and will help to understand their roles in ecology and human health. Recently, droplet microfluidics is emerging as a promising technology for microbial studies with value in microbial cultivating, screening, and sequencing. This review aims to provide an overview of droplet microfluidics techniques for microbial research. First, some critical points or steps in the microfluidic system are introduced, such as droplet stabilization, manipulation, and detection. We then highlight the recent progress of droplet-based methods for microbiological applications, from high-throughput single-cell cultivation, screening to the targeted or whole-genome sequencing of single cells.

show abstract

“…Reducing surfactant concentrations or maintaining droplet‐to‐droplet distance can limit exchange, although this not always applicable, especially when long‐term incubation is required. In fact, reported studies on the exchange kinetics of some fluorescent dyes have shown that retention times could range from days (e.g., fluorescein ) to seconds to minutes (e.g., rhodamine , coumarin and resorufin ). It has been suggested that increasing hydrophilicity of the dye by introducing more polar groups can reduce micellar transport of molecule .…”

Section: Droplet‐based Microfluidic Technologies For De Of Enzymesmentioning

confidence: 99%

High‐throughput droplet‐based microfluidics for directed evolution of enzymes

Chiu

Stavrakis

2019

Electrophoresis

View full text Add to dashboard Cite

Natural enzymes have evolved over millions of years to allow for their effective operation within specific environments. However, it is significant to note that despite their wide structural and chemical diversity, relatively few natural enzymes have been successfully applied to industrial processes. To address this limitation, directed evolution (DE) (a method that mimics the process of natural selection to evolve proteins toward a user-defined goal) coupled with droplet-based microfluidics allows the detailed analysis of millions of enzyme variants on ultra-short timescales, and thus the design of novel enzymes with bespoke properties. In this review, we aim at presenting the development of DE over the last years and highlighting the most important advancements in droplet-based microfluidics, made in this context towards the high-throughput demands of enzyme optimization. Specifically, an overview of the range of microfluidic unit operations available for the construction of DE platforms is provided, focusing on their suitability and benefits for cell-based assays, as in the case of directed evolution experimentations.

show abstract

Synthesis of new hydrophilic rhodamine based enzymatic substrates compatible with droplet-based microfluidic assays

Abstract: Here we report the conception, synthesis and evaluation of new hydrophilic rhodamine-based enzymatic substrates for detection of peptidase activity compatible with high-throughput screening using droplet-based microfluidics.

Cited by 24 publications

References 35 publications

Controlled droplet microfluidic systems for multistep chemical and biological assays

Controlled droplet microfluidic systems for multistep chemical and biological assays

One cell at a time: droplet-based microbial cultivation, screening and sequencing

High‐throughput droplet‐based microfluidics for directed evolution of enzymes

Contact Info

Product

Resources

About