Concept

The concept of the device T shares the first steps with the device C (Sec. 4.1):

Protein and multivalent ions are encapsulated as microfluidic droplets. Shortly before the drops are formed, protein and multivalent ions get in contact for the first time. The content is mixed quickly. Then an individual drop is trapped and its content is imaged for several minutes.

A key feature of this device is that it provides imaging of the interaction of protein and the multivalent ions rapidly after the first protein-salt contact at time t_init.

‘Rapidly’ means that data are recorded already after 1-5 s. Rapid content imaging could also be achieved faster by following drops along a serpentine channel. [118]

However, the details of filaments in the moving drops are smeared out as the drops move through the channel. Our approach provides both rapid imaging and good imaging results of the drop content over time. This combination of a rapid imaging start and a trapped drop is the main benefit of this microfluidic concept.

4 MICROFLUIDIC DEVICES FOR PROTEIN STUDIES

4.2.2 Device design and function

The microfluidic device design to image the drop content rapidly after drop pro-duction is shown in Fig. 4.4. Aqueous drops are produced in a flow focusing geometry. [7] Similar to the device C, the aqueous phase is composed of three aqueous components aq 1, aq 2 and aq 3 that join up into one channel shortly before the aqueous phase is dispersed (Fig. 4.4b). [118] The aq 3 component is supplied using the method of staggered fluids (Sec 5.3). After their production, the drops pass a 2.5 mm long and straight channel (Fig. 4.4a and c). In this chan-nel, the drops are plug-like and – like the serpentine channel in the device C –

outlet

2 mm inlets outlet high oil counter flow^55µm

drop

Figure 4.4: Concept of the device T. The aim of the device T is to image individual drops containing vimentin and a multivalent salt over time and rapidly after the rst contact of the multivalent salt and the protein. b) The three aqueous components get into contact shortly before they are encapsulated as drops. [118] c) Ecient mixing is achieved as the plug-like drops move through a straight channel. [27] d) In `U'-shaped traps [58], the content of drops is imaged over time. The trap region is at because of a step in the channel. The reduced channel height improves the stability of drop trapping. To empty the traps, a higher counter oil ow can be applied. This allows for serial drop imaging.

(Adapted from reference [35] with permission from The Royal Society of Chemistry)

Device T : Time-lapse studies 4.2

procedure u_{top oil} u_{bottom oil} u_{aq 1} u_{aq 2} u_{aq 3} device startup for≈1h 2.4 4.8 1.2 1.2 2.4 drop trapping and imaging 2.4 24 1.2 1.2 1.2 drop release for≈10s 2.4 ≈48 1.2 1.2 1.2

Table 4.2: Flow velocities. Typical ow velocities u in mm s⁻¹ applied during the dierent procedures of the device T operation. They apply for imaging of drops which have a diameter of 100µm.

it used to achieve fast mixing. [27] In contrast to the device C, the drops within the device T have a larger volume at the flow speeds we typically use (Tab. 4.2).

Therefore, their plug-length would span over several serpentine windings, if they passed a serpentine channel as in the device C (Fig. 4.1d). The fast moving drops would most likely break up into smaller drops. Therefore, we use a mixing chan-nel in the device T that is straight (Fig. 4.4c). Like in the case of the serpentine channel, fast mixing of the drop content is achieved. By calculation the mixing should be finished within two seconds after drop production (Sec. 2.3.3; calcula-tion in Sec. 5.1).

After the straight channel, the drops enter a broadened region where they are densified as most of the oil and excess drops leave the device via two lateral out-let channels (Fig. 4.4d). In the center of the broadened region the drops face a step in the channel. At this step the channel height is reduced from 33 µm to 18 µm. The drops accumulate in front of this barrier and they are continuously exchanged by newly arriving drops. The adjoining flat region has several ‘U’-shaped traps. [58] When the oil counter flow from the bottom is relatively low (Fig. 4.4a and d), drops enter the flat region and flow into the traps. In this po-sition, the drop content is imaged over time. The dwell time of the drops in the traps can be more than 10 min. However, this time shows large variation (Sec.

4.2.3) due to the small slit at the lower end of the traps. During drop content imaging, the flow in the device is not stopped. As a result the drops are some-times squeezed through this slit. At the same time, the slits are needed to empty the traps by an increase in the bottom oil flow (Fig. 4.4d). This allows for a serial imaging of the drop content. A further improvement of the device T could be an adjustment of the size of the slit. A smaller slit would increase the time in the drop. At the same time, the slit has to be large enough to ensure trapping and release of the drops.

The first row of the trap region is the most important one, as it is the fastest row

4 MICROFLUIDIC DEVICES FOR PROTEIN STUDIES

300 µm

flathighhigh

step step

step

Figure 4.5: Multiple-sized drop traps. A device T that can trap drops with a diameter of 55 µm up to 250 µm. The drops are imaged in the traps that t best to the drop diameter. Drops that do not t the trap size are ignored during imaging. (left: lithography master, right: corresponding PDMS device)

reached. The time that passes between the first contact of the aqueous compo-nents and the first image acquisition of a drop content is about 1-5 s (Sec. 4.2.3).

The flat region in the device is essential. First, it renders the drop trapping more stable. Second, it reduces the number of incoming drops that could disturb drop content imaging. We find that the trapping mechanism is not restricted to a cer-tain drop size. Complementary to the usage of one trap size as given in Fig. 4.4, a design with traps of different sizes works, too. This way it can be decided in each experiment, which drop size is of interest (Fig. 4.5). For drops of about 100µm in diameter (flat region), we give typical flow speeds in the device in Tab. 4.2.