This protocol describe our method to fabricate and operate on pneumatically-driven microfluidic platform for multi-particle concentrations that overcomes shortcomings such as particle clogging and complex structures. This methods makes it possible to process an unlimited number of particles, concentrate on a large number of small particles, and prevent unwanted cell damage, and increases entropy efficiency. Due to the importance of biological analysis, microfluidic and biomedical microelectro mechanical systems technologies are used to develop and study devices for the purification and collection of micro-materials.
To begin, use a pre-prepared pneumatic valve channel SU8 mold to replicate the PDMS layer for pneumatically controlling the valve. Pour 10 milliliters of liquid PDMS and one milliliter of curing agent into a prepared pneumatic valve channel mold, and heat activate at 90 degrees Celsius for 30 minutes. After the PDMS structures are cured, separate the SU8 mold.
Punch three 1.5 millimeter pneumatic ports into the pneumatic valve channel using a 1.5 millimeter puncture. Pour 10 milliliters of liquid PDMS and one milliliter of curing agent into a clean SU8 mold. Spin coat for 15 seconds at 1500 revolutions per minute using a spin coater, then heat activate at 90 degrees Celsius for 30 minutes.
Separate the SU8 mold after the PDMS structures are cured. Treat the PDMS structure with atmospheric plasma for 20 seconds. Using a microscope, align the plasma treated PDMS structures according to the channel structure.
Bond the aligned PDMS structures by heating them at 90 degrees Celsius for 30 minutes. Using a 1.5 millimeter puncture, make a 1.5 millimeter diameter hole in the fluid channel inlet and outlets within the pneumatic channel part bonded to the thin diaphragm layer. Replicate both sides of the PDMS layer using two SU8 molds to make a microfluidic channel.
Use a curved and rectangular microfluidic channel mold on the front, and a microfluidic interconnection channel mold on the rear. Pour 10 milliliters of liquid PDMS and one milliliter of curing agent into the curved and rectangular microfluidic channel mold, and spin coat it at 1200 revolutions per minute for 15 seconds, then create molds for the curved fluid chamber and fluid channels by heat activation at 90 degrees Celsius for 30 minutes. Separate the PDMS layer on which the microfluidic channel is formed.
Then treat it with atmospheric plasma for 20 seconds to make a heat-activated mold covering the sealed vent wall by bonding to the glass wafer. Pour three milliliters of liquid PDMS into the interconnection channel of the SU8 mold. Arrange the structure, fabricated with the interconnection channel mold, in liquid PDMS on the microfluidic interconnect channel mold.
Then dry the superimposed structure at 130 degrees Celsius for 30 minutes. After curing, remove the front SU8 mold from the microfluidic channel network layer and carefully peel off the rear PDMS mold. Pour 10 milliliters of liquid PDMS and one milliliter of curing agent into a clean SU8 mold and heat activate it at 90 degrees Celsius for 30 minutes.
Separate the SU8 mold after the PDMS structures are cured. Treat the PDMS microfluidic interconnection channel molds with atmospheric plasma for 20 seconds. Using a microscope, align the plasma-treated PDMS structures according to the channel structure.
Bond the aligned PDMS structures by heating at 90 degrees Celsius for 30 minutes. Align the PDMS structures prepared during this process according to the channel structure and bond them by treating with atmospheric plasma for 20 seconds. Using a 10 milliliter syringe, fill the microfluidic channel with bubble-free, demineralized water.
To control the pressure of working fluid and the three pneumatic valves that control the microbead flow, insert a precision pressure controller with four or more outlet channels for the working fluid into the microfluidic platform. Prepare carboxyl polystyrene test particles of various sizes in distilled water. To control the flow rate of the working fluid, fill half of a glass bottle with the water and connect the glass bottle cap to the controller output channel and microvalve.
Using an inverted microscope, observe all the platform operations and measure the operating flow rate over time at the outlet by a liquid flow meter. Inject the particle or fluid mixture under pressure at the inlet with the particle valve. Apply pressure to the CIV valve at 15 kilopascals, and particle valve at 18 kilopascals to actuate the valve.
When the particles are concentrated, apply pressure only to the fluid valve. The flow rate of the fluids was divided into a four-stage platform operation. The first stage was the loading state.
The working fluid and particles were almost identical as the microfluidic channel network exhibited structural symmetry. The second stage was the blocking state. The flow path narrowed, and the flow rate measured at the outlet port was reduced by hydraulic resistance.
The third stage was the concentration state. The measured QP was close to zero, and the QF was about 1.42 times that of the blocking state. The final stage was the release state.
The resulting flow and concentration rates proved that the sequential actuation programmed with the pneumatic valve work well due to flow changes. Particles were concentrated and accumulated in the collection area when the CIV valve and particle valve were closed, and all collected, concentrated particles were released within four seconds when only the fluid valve was closed. An essential part of this procedure is curing the rear structure where the PDMS layer is implanted by the some more pressure of the air layer.
And the deformed film layer is suddenly activated. This platform can be used for auto-pretreatment of very concentrated and straight and suspended bioparticles, as the operation is not affected by the properties of the physical particles.
