Abstract
Microfluidic single-cell cultivation (MSCC) is an emerging field within fundamental as well as applied biology. During the last years, most MSCCs were performed at constant environmental conditions. Recently, MSCC at oscillating and dynamic environmental conditions has started to gain significant interest in the research community for the investigation of cellular behavior. Herein, an overview of this topic is given and microfluidic concepts that enable oscillating and dynamic control of environmental conditions with a focus on medium conditions are discussed, and their application in single-cell research for the cultivation of both mammalian and microbial cell systems is demonstrated. Furthermore, perspectives for performing MSCC at complex dynamic environmental profiles of single parameters and multiparameters (e.g., pH and O2 ) in amplitude and time are discussed. The technical progress in this field provides completely new experimental approaches and lays the foundation for systematic analysis of cellular metabolism at fluctuating environments.
Highlights
Introduction small microcolonies with up to1000 cells[4,5] and cell growth is restricted to a monolayer
We propose to differentiate between five different categories on how dynamically controlled environments (DCE) can be created in microfluidic devices (Figure 2F)
Depending on the flow rate for example 1 μL s−1, a temporal resolution of around 10 s can be experimentally realized with a microfluidic channel with a height of ≈100 μm and a volume of ≈0.6 μL cm−1.[67,68] The DCE can oscillate in a range of seconds so that different feed streams can be feed in short time-scales
Summary
Environmental conditions in natural habitats of bacteria are generally not constant but vary in concentration over the time.[26,27] In their natural habitat (e.g., soil and ocean) and artificial habitats (e.g., bioreactors) microbial cells are exposed to changing environmental stimuli. The perfusion of media during conventional MSCC leads to continuous medium supply and the removal of products and by-products and constant environment conditions over the cultivation time (Figure 2C) In their natural habitat cells are exposed to randomly changing environmental conditions with changes of frequency and amplitude of environmental parameters (Figure 2E).[38] This includes gradients in nutrients,[39] temperature,[40] pressure,[41] pH value,[42] and oxygen level.[43] Even in simulated natural habitats such as laboratory bioreactors, cells are exposed to short-term environmental fluctuations.[26,44] Here, the fluctuations can be smaller in amplitude but can be faster in frequency.[45] To analyze the influence of environmental fluctuation in laboratory scale as well as natural habitats novel microfluidic systems must be developed to mimic fluctuating environmental conditions (Figure 2D).[46]. An environmental profile can be established with the same amplitude and width, only the frequency between the pulses (f1 and f2) is varied
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