Combining Microfluidics and Microrheology to Determine Rheological Properties of Soft Matter during Repeated Phase Transitions

The overall goal of this procedure is to use microfluidics to induce consecutive phase transitions in a single soft matter sample by exchanging the surrounding fluid while simultaneously measuring changes in the rheological properties using multiple particle tracking microrheology. This method can answer key questions in the soft matter field, such as the impact of phase transitions, including molecular or colloidal rearrangement on dynamic rheological properties. This technique's main advantage is that the sample remains in place during multiple phase transitions, enabling real-time characterization.

This allows quantification of rheological properties, especially unique properties in heterogeneous systems. To begin the procedure, print the microfluidic device negative on a clear acetate sheet, and warm up a high-intensity UV lamp. On a second clear acetate sheet, trace the corners of a 75 millimeter by 50 millimeter slide.

Place one millimeter high glass spacers on the corners. Pour about five milliliters of UV-curable thiol-ene resin in the center of the spacers. Carefully lay a 75 millimeter by 50 millimeter by one millimeter glass slide on the spacers to cover the resin.

Ensure that there are no air bubbles between the slide and the resin. Place the printed negative on the glass slide. Carefully drag the assembly under the UV lamp, and allow the resin to cure for 45 seconds.

Then, slide the assembly away from the UV lamp, and remove the patterned glass slide or microfluidic stamp. Immerse the microfluidic stamp in acetone for no more than 10 seconds, and then immerse the stamp in ethanol for the same length of time. Repeat this immersion sequence once.

Then, submerge the stamp in distilled water. Working with minimal hand movement, use a cotton swab to remove the remaining unreacted resin without touching the cured resin. Replace the cotton swab when it becomes saturated with water.

Cure the stamp under UV light for another 30 minutes. Then, mix 70 grams of PDMS base with cross-linking agent in a one-to-10 ratio, and degas the PDMS for one hour or until the PDMS is no longer turbid. Place the microfluidic stamp face up in a 150 millimeter plastic Petri dish.

Slowly pour the PDMS into the dish, pouring close to the dish surface to minimize bubble formation, to completely cover the stamp. Cover the Petri dish, and cure the PDMS at 55 degrees Celsius overnight. Use a knife to cut away the excess PDMS.

Separate the patterned PDMS from the stamp by hand. Remove the stamp and the excess PDMS from the Petri dish. Then, place the dish on a sheet of paper printed with the locations of the symmetric outer holes for the sample chamber.

Align the patterned PDMS with the guide, and use a 5 millimeter biopsy punch to punch holes through the PDMS at the marked locations. The key feature of this device is the symmetry between inlet channels and the same chamber. This enables exchange of fluid without loss of sample.

It is important to use a guide pattern to ensure accuracy when cutting the channels. Lastly, punch a hole in the center of the sample chamber, at the inner edge of the suction chamber, and at each end of the long outer channels. Cover the Petri dish when storing the patterned PDMS.

One day before the device assembly, prepare the preconverted fluid that will be used to create glass walls in the microfluidic device. To begin the device assembly, plasma treat the patterned PDMS and a thin glass slide for 40 seconds. Carefully press together the plasma-treated faces of the slide and the patterned PDMS.

Apply UV-curable resin to the seam between the PDMS and the slide, and cure the resin under low-intensity UV light for five minutes. Next, insert thermoplastic tubing fitted with stainless steel connectors into all but one corner hole of the patterned PDMS. Connect a three milliliter syringe of preconverted fluid to the last hole using an 18 gauge needle, thermoplastic tubing, and a connector.

Fill the microfluidic device with preconverted fluid. Then, place the device on a hot plate heated to 100 degrees Celsius, and, over the course of 10 seconds, flow three milliliters of preconverted fluid through the device to fabricate the glass walls. Then, remove the device from the hot plate.

Use a 30 milliliter syringe of air to expel excess preconverted fluid. Slowly flow 15 milliliters of chloroform through the device, followed by 30 milliliters of ethanol, each over the course of about one minute. Then, push air through the microfluidic device until dry.

Remove the tubing from the holes. Next, use UV-curable resin to attach one millimeter thick glass strips to the undersides of the long edges of the device. Cut a 30 by 30 millimeter piece of excess cured PDMS, and punch a hole wider than the sample chamber in the center to form the solvent basin.

Plasma treat the basin and the microfluidic device for 40 seconds. Place the plasma-treated face of the basin on the device with the sample chamber centered in the hole. Apply gentle pressure to fix the basin to the device.

Lastly, cut six five millimeter squares from the excess PDMS. Use a 5 millimeter biopsy punch to cut a hole halfway through each square. Insert a stainless steel connector into each hole to form the PDMS stoppers.

After loading the soft matter sample into a one milliliter syringe, insert thermoplastic tubing into three of the device's corner inlet channels and the suction chamber outlet. Fill another syringe with deionized water. Attach thermoplastic tubing to the syringe, and expel any air bubbles.

Connect the syringe to the fourth corner inlet channel. Fill the device and the solvent basin with deionized water. Ensure that there are no bubbles in the sample chamber or in the microfluidic channels.

Top off the water in the solvent basin with a transfer pipette if needed. It is essential that the microfluidic device is set up correctly. Most importantly, ensure that there are no air bubbles in the device, as they can disrupt both the microfluidic flow and the microrheology data.

Then, inject 10 microliters of the soft matter sample into the sample chamber via the center inlet at about two microliters per second. Stopper the center inlet after loading the sample. Using an inverted microscope and a high-speed camera, record at least 20 videos of the Brownian motion of the probes in the sample mixture over time intervals appropriate for the duration of the phase transition.

For gelation, continue recording videos until probe movement has stopped completely. For degradation, continue recording videos until the probes are completely diffusive. Then, remove the device from the microscope stage.

To exchange the water for a higher density fluid, withdraw the water from the solvent basin using a transfer pipette, and pipette four milliliters of the higher density fluid into the basin. Return the device to the microscope stage, and continue recording videos. To exchange a higher density fluid for water or another lower density fluid, connect a syringe to the suction chamber channel via an 18 gauge needle and thermoplastic tubing.

Mount the syringe on a syringe pump set to withdraw at one milliliter per minute. Pipette most of the high-density fluid from the solvent basin, and rinse the basin with the low-density fluid three times, never allowing the basin to drain completely. Fill the basin with low-density fluid after rinsing.

Fill a transfer pipette with the low-density fluid. Then, use the syringe pump to withdraw the high-density fluid for one minute. Add the low-density fluid dropwise to the basin as needed during the fluid exchange.

Then, stopper the suction chamber, and return the device to the microscope stage for recording. Acquire videos of degradation gelation cycles in this way for the desired number of cycles or until there are insufficient probes for measurement. The ensemble averaged mean-squared displacement of probe particle trajectories was tracked during successive phase transitions of a hydrogenated castor oil system between gel and sol.

Gelation was induced by gravity flow exchange of water for a higher density gelling agent. Degradation was induced by suction flow exchange of the gelling agent for water. The sample chamber was reflushed with water when the phase behavior suggested that the gelling agent had not been fully removed from the chamber.

No changes to the rheological properties were observed when the sample was left in the sol phase with no fluid exchange for about 600 minutes. Repeated phase transitions did not induce observable changes in equilibrium rheological properties, as shown by the logarithmic slopes of the MSD curves or alpha returning to the same values for each phase. The equilibrium alpha value of the sol phase was less than 1.0, indicating that the material retained some structure even after degradation.

The gel underwent an increase in structural heterogeneity during the gel to sol phase transition, as indicated by the non-Gaussian parameter peaks during degradation cycles. Colloidal rearrangement could occur immediately after fluid exchange, as indicated by increases in alpha. We first had the idea for this method to determine whether phase transitions were causing permanent changes in rheological properties and microstructure of a soft material.

In the HCO system, this device enabled measurement of two possible mechanisms for these permanent changes, colloidal rearrangement during phase transitions or shear induced during sample preparation. Once mastered, this technique can be used to identify the rheological properties of a soft matter system over many phase transitions in response to changes in the surrounding fluid environment.