Microfluidic device to study patterned SVs under a switch-

This project is in collaboration with the group of Prof. Reinhard Jahn from the Laboratory of Neurobiology of the Max Plank Institute for Biophysical Chemistry in Göttingen. To measure the interaction of molecules with printed SVs in flow, a three-inlet device (channel length 2.3 cm, width 1 mm and height 100µm) is

Figure 3.4: Sketch of the construction of the five-inlet microfluidic device to study protein aggregation. The central channel of the microfluidic device has a constriction to reduce the height. One side of the constriction is already integrated in the PDMS replica of master 1. The second side of the constriction is created directly on the glass coverslip. a) A cut-open PDMS replica of master 2 is applied to a clean glass slide. b) A drop of liquid adhesive (NOA H83) fills, via capillary forces, the channel. c) After being cured by UV light for three hours, the liquid adhesive channel is furthermore cured under UV light without the PDMS slab. d) The cured glue channel is aligned with the PDMS replica of master 1 and the two sides are bound after air plasma treatment. e) The device is ready to be employed in experiments.

employed. This microfluidic device is used to flush different fluids on top of the SV pattern in a controlled manner. To measure how fast, for example, the SVs react to a different pH or to a particular molecule, we have to be able to change the reactant agent quickly. The idea is to have a device with the SV pattern in the center of the outlet channel and to control the flow in a way to be able to switch the solution flowing, for example, from the one contained in inlet 1 to the one in inlet 2, in a controlled and fast manner, as shown in Figure 3.5.

To achieve the switching of the solution flowing on top of the patterned SV, a rectangular function is applied to the side inlets of the microfluidic device, with a maximum velocity of 1 mm/s, a minimum velocity of 0.1 mm/s and width 10s, as shown in Figure 4.38a (Section 4.2.3). When the flow from the side-1 channel has the maximum velocity, as in Figure 3.5a, the flow from the side-2 channel has the minimum velocity. In the central outlet, the flow from side-1 pushes the solution from side-2 towards the channel wall, filling the central part of the outlet channel where the SV are patterned. When the flow from the side-1 channel has the minimum velocity, as in Figure 3.5b, the flow from the side-2 channel has the maximum velocity. In this case, the solution from side-1 channel is pushed away from the central part of the outlet by the solution from side-2 channel. Thus, the patterned SVs are now exposed to the solution coming from the side-2 inlet.

The master wafers are created in the 1000 class clean room of the physics faculty at the University of Göttingen by standard photo-lithography methods [115, 116]. Briefly, a 100µm layer of negative photo-resist (SU8-3050) is

uni-Figure 3.5: Schematic representation of the microfluidic device to study SVs under a switchable flow. When the velocity is maximum in one side inlet, for example side 1, the solution from that specific inlet fills the central outlet channel. When the velocity from that side inlet reaches the minimum, the solution from that inlet is not anymore in the center of the device, but the solution from the second side inlet fills the outlet channel.

formly spin coated on top of a 2-inch cleaned silicon wafer with velocities accord-ing to Table 3.1. The wafers are soft baked at 95^◦C for 45 minutes and then exposed to UV light through a photomask via the mask aligner. The wafers are baked again for 15 minutes at 95^◦C and developed for 15 minutes. After cleaning the wafers with isopropanol, they are coated overnight with (heptafluropropyl)-trimethylsilane. PDMS replicas are created by mixing the liquid PDMS with the cross-linker at a ratio of 10:1 and pouring the mixture on the wafers. After remov-ing the air-bubbles with a desiccator, the PDMS is cured in the oven (DryLine, VWR, Darmstadt, Germany) at 65 ^◦C for 1 hour.

As in this case, SVs are patterned on the glass surface, which is the bottom of the microfluidic device, plasma treatment cannot be used to covalently bind the PDMS channel replica with the glass coverslip because the plasma would destroy the previously patterned SVs on the surface. To assemble the device, the PDMS channel and the patterned glass coverslips are placed inside an home-made device holder (a sketch is presented in Figure 3.6) inspired by [118, 134]. The device holder consists of two metal plates clamped together with 8 bolt screws.

The patterned glass coverslip is inserted between the two plates. A window in the bottom metal plate allows us to image the patterns with a microscope. The PDMS replica with the channel is aligned on top of the glass coverslip effectively closing the device. Uniform pressure on the device is achieve by inserting a plastic (PVC), or glass, plate (1 mm of thickness) before the second metal plate. The uniform pressure prevents also leaking of the device. Tubings are inserted to the device through holes in the top metal plate and in the plastic plate. A photograph of the microfluidic device mounted in the holder is shown in Figure 3.6b.

Figure 3.6: Device holder for microfluidic chips with patterned molecules on the glass coverslip.

The microfluidic device is held together by two metal plates screwed together via 8 bolts screws.

The bottom metal plate has an opening to allow us to image the microfluidic device in an inverted microscope. The PDMS replica of the channels is mounted on top of the patterned glass slide closing the microfluidic device. On top of the PDMS, a thick glass slide or a PVC plate is used to uniformly spread the pressure on the device preventing leakage. Tubings are inserted via holes on the top metal plate and the PVC plate.