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Abstract:
The interior as well as the exterior of cells is governed by networks composed of fi-
brous proteins. The mesh size of these networks is on the order of micrometers and
therefore distinguishes microfluidics as an excellent tool to gain insight into principal
mechanisms of single molecule behavior, on the one hand, and the interplay
and self-assembly of the network constituents, on the other hand. Here, we present
new results derived from biomimetic investigations of two different systems, namely
single molecule experiments on actin, one of the most important intracellular proteins,
and in situ observation of the fibril formation of collagen I, the most abundant
extracellular protein. The use of microfluidic channels fabricated by means of soft
photolithography as the principle tool for our experiments enables us to manipulate
the molecules via confining wall potentials and hydrodynamic flow fields, analyze
their mechanical behavior, and observe time and spatially resolved reactions. Furthermore,
microfluidics is very well suited for combination with different observation
methods such as fluorescence microscopy, polarized light microscopy, and X-ray microdiffraction.
Analyzing single fluctuating actin filaments under the influence of confinement yields
a thorough characterization of the mechanics of the system. The biomacromolecules
are observed by means of fluorescence microscopy. We find that the behavior of the
biopolymers depends on their contour length L and the influence of the microfluidic
environment. The confining energy is considered as a parabolic wall potential. Thus,
we succeed to remarkably well describe the competition between bending energy and
confining energy. Moreover, the results are consistent with Monte Carlo simulations
and with scaling laws for the deflection length λ and the segment distribution in the
channels.
The experiments on collagen I give insight into the dynamic evolution of the hierarchical
organization of native collagen fibrils. We use a hydrodynamic focusing
and diffusive mixing device to establish a stable pH-gradient within the microfluidic
channels. Therefore, we are able to perform non-equilibrium measurements in the
laminar flow and observe different stages of the self-assembly process at different positions within the same system. We characterize the system on a microscopic length
scale by availing the birefringent properties of collagen. Additionally, using X-ray
microdiffraction the dynamic formation of critical subunits of collagen fibrils can
be observed. Furthermore, we demonstrate that finite element method simulations
provide a good description of our experimental results regarding diffusive phenomena,
influence of the solution viscosity on the flow profile, and pH distribution.
The experiments presented here elucidate the principle understanding of the studied
biological systems and furthermore show the ability of microfluidic tools to advance
the diverse field of life science.
