KEYWORDS: Bacteria, Capillaries, Two photon polymerization, 3D microstructuring, Optical tweezers, Optical micromanipulation, Current controlled current source
Swimming bacteria exploit viscous drag forces to generate propulsion in low Reynolds number environments. A rotating helical flagellar bundle can propel the cell body at typical speeds of ten body lengths per second. Not surprisingly, this ability to efficiently swim is preserved even in confining micro-environments which constitute their typical habitat. Quantitative studies would require the ability of fabricating complex environments with controlled geometrical properties. Experimental studies so far were limited to large diameter micro capillaries or 2D confinement. In this last case, E.coli has been shown to swim with an unaltered speed even when the gap size is slightly larger than the cell body thickness. The case of tight 1D confinement is however more challenging requiring 3D fabrication capabilities. Using two-photon polymerization we fabricate 3D microstructures that can confine swimming bacteria in quasi 1D geometries.
We observe individual E.coli cells swimming through a sequence of micro-tunnels with progressively decreasing diameters. We demonstrate that E.coli motility is preserved also in tight 1D confinement. Moreover we find that there's an optimal channel diameter for which the increase in flagellar thrust due to 1D confinement can even overcome the increased drag on the cell body resulting in swimming speeds that can be up to 15% larger then the bulk speed.
Lorenz-Mie scattering theory allows to predict the field scattered by spherical objects illuminated by coherent light. By fitting the fringe pattern resulting from the interference of incident and scattered light, it is possible to track and size colloidal particles with a few nanometer precision.
Using digital holographic microscopy (DHM) we extend the applications of Lorenz-Mie theory to hollow spherical structures and to extremely high pressure conditions.
On the one hand, we geometrically and optically characterize complex colloids as polymer-shelled microbubbles, with high precision, low costs and short acquisition time. These microbubbles are likely to be unique tools for targeted drug delivery and are currently used as contrast agents for sonography. We measured size, shell thickness and refractive index for hundreds of polymeric microbubbles showing that shell thickness displays a large variation that is strongly correlated with its refractive index and thus with its composition.
On the other hand we demonstrate that DHM can be used for accurate 3D tracking and sizing of a holographically trapped colloidal probe in a diamond anvil cell (DAC). Polystyrene beads were trapped in water up to Gigapascal pressures while simultaneously recording in-line holograms at 1 KHz frame rate. This technique may potentially provide a new method for spatially resolved pressure measurements inside a DAC.
Diamond anvil cells can be used to study the behavior of materials at high pressure by compressing small samples
up to hundreds of GigaPascals. There is no mechanical access to the sample once the cell is pressurized but it
is possible to observe the sample through the diamond windows. Optical tweezers can be used to measure the
mechanical properties of fluids, such as viscosity, by trapping and monitoring micron sized spheres suspended in
the fluid. We use a diamond anvil cell within a modified optical tweezers instrument to measure the viscosity
of water as a function of pressure up to 1:3GPa. Development of this technique will allow investigations of the
mechanical changes in biological cells and other soft materials placed under high pressure.
Diamond anvil cells allow us to study the behaviour of materials at pressures up to hundreds of gigaPascals in a small and convenient instrument, however physical access to the sample is impossible once it is pressurised. Optical tweezers use tightly focussed lasers to trap and hold microscopic objects, and their ability to measure nanometric displacements and femtonewton forces makes them ubiquitous across the nano and bio sciences. We show that optical tweezers can be used to hold and manipulate particles in such a cell, in the ``macro tweezers'' geometry allowing us to use objective lenses with a higher working distance. Traps are structured to overcome the limitations imposed by the sample cell. Wedemonstrate the effectiveness of the technique by measuring water's viscosity up to 1.2 GPa. The maximum pressure reached was limited by the water crystallising under pressure.
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