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Journal Articles Analytical Chemistry Year : 2015

Measuring Fast and Slow Enzyme Kinetics in Stationary Droplets

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Abstract

We present a new microfluidic platform for the study of enzymtatic reactions using static droplets on demand. This allows us to monitor both fast and slow reactions with the same device and minute amounts of reagents. The droplets are produced and displaced using confinement gradients, which allows the experiments to be performed without having any mean flow of the external phase. Our device is used to produce six different pairs of drops, which are placed side by side in the same microfluidic chamber. A laser pulse is then used to trigger the fusion of each pair, thus initiating a chemcial reaction. Imaging is used to monitor the time evolution of enzymatic reactions. In the case of slow reactions, the reagents are completely mixed before any reaction is detected. This allows us to use standard Michaelis–Menten theory to analyze the time evolution. In the case of fast reactions, the time evolution takes place through a reaction-diffusion process, for which we develop a model that incorporates enzymatic reactions in the reaction terms. The theoretical predictions from this model are then compared to experiments in order to provide measurements of the chemical kinetics. The approach of producing droplets through confinement gradients and analyzing reactions within stationary drops provides an ultralow consumption platform. The physical principles are simple and robust, which suggests that the platform can be automated to reach large throughput analyses of enzymes.
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Dates and versions

hal-01255747 , version 1 (13-01-2016)

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Etienne Fradet, Christopher Bayer, Florian Hollfelder, Charles N. Baroud. Measuring Fast and Slow Enzyme Kinetics in Stationary Droplets. Analytical Chemistry, 2015, 87 (23), pp.11915-11922. ⟨10.1021/acs.analchem.5b03567⟩. ⟨hal-01255747⟩
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