MICROFLUIDICS RESULTS:

Our ultimate goal of making cells battle within a microfluidic chip for entertainment purposes is a project which has never been attempted.Therefore, the unique design needed for our microfluidics chips had to be created from scratch. The design process required the operation of AutoCAD: a software application for drafting 2D and 3D design. Since microfluidic chips are normally used for biological assays, we had to take into consideration the forces acting on a droplet with isolated cells, and how the droplets can be manipulated to battle each other. Over the course of two months, the team learned how to use the program to design a various array of microfluidic chips ranging from an intricate mosaic of channels to creatively ingenious designs. The team payed attention to detail, from the measurements of the channel width, outlets, and inlets to accommodate E.coli and S.cerevisiae: with all these limitations the team produced eight different chip designs. In addition, to acheive a “head-on” a battle the mechanics of the droplet had to be manipulated, therefore electrodes were added to induce a turbulent flow when the droplets merge. The next step is fabricating the master microfluidics chip through a process called photolithography where a silicon wafer that is coated with a thin layer of of SU-8 photoresist is exposed to UV-light and the photomask pattern is transferred to the silicon wafer.

The first step in microfluidic chip fabrication is making a master--this requires a process known as photolithography which is performed in a cleanroom since photolithography requires an extremely clean substrate. Photolithography is employed mainly because integrated circuits can be made through optical erosion, in contrast to physically cutting into the integrated circuit. Therefore, photolithography was used to fabricate the chip because of its precise incisions. Photolithography begins with cleaning the substrate by rinsing the substrate.The second phase of cleaning the substrate is purging the substrate of all liquids, and then it heated. The second step in fabricating the master is placing the clean substrate onto the spin-coater and then a drop of HMDS is applied to the substrate: this facilitates and increases adhesion of the SU-8 photoresist to the substrate.The substrate is then spinned to form a thin layer of HDMs, afterwards SU-8 photoresist is poured onto the center of the substrate and spinned.This step was a crucial to form a uniform and air-bubble free spin-coated layer of SU-8 photoresist. The wafer was then “soft-baked”, the spin coated wafer was placed on a leveled hot plate. Subsequently, the wafer is exposed to a UV light under a photomask, as a result the ultraviolet light causes the exposed areas of the substrate to dissolve and a pattern is created.


Following exposure, the wafer submerged into it a developer causing the photo-masked areas of the wafer to be washed away, while the exposed part remained and the pattern of interest was embossed. After development, the wafer is hard baked and the wafer is hardened.This is what is known as a master, since this wafer can be now be used to generate an infinite number of chips.

PDMS is poured over the silicon master mold, cured and then peeled off; holes are then punched for inlets and outlets. After plasma exposure, PDMS is bonded to a cover glass to yield the final microfluidic device. In addition, by pouring PDMS, a transparent bio-compatible polymer often used in creating microfluidic chips, on top of the master and baked overnight--a solid slab of PDMS which has been etched with our designed patterns was produced. The height of the channels is dependent on the height of the remaining photoresist on the master. The etched side of the PDMS is then bonded onto an appropriate flat surface such as the glass surface of a microscope slide. The space between the microscope slide and the etched parts of the PDMS will create the desired channels. Holes are punched into the PDMS slab wherever we desire to have an inlet or an outlet.

Anders S Hansen, Nan Hao & Erin K O'Shea. High-throughput Microfluidics to Control and Measure Signaling Dynamics in Single Yeast Cells. Digital image. Http://www.nature.coml. N.p., n.d. Web. 19 Oct. 2016.

Once the chip is made, the channels must be coated with a hydrophobic layer this is done by coating the channels with Pico-glide a surface coating agent to improve droplet performance and stability.. The hydrophobic layer is Pico-glide made from a fluorous polyether polymer that chemically reacts with etched glass and PDMS surfaces. When Pico-glide is flowed through the channels a uniform layer will covalently bond to the microfluidic channel surfaces of PDMS and glass microfluidic devices.

In order to flow fluids through our chip, specifically: cell suspensions, oil phase, chemicals, reagents are inserted into a special syringe with a gauge needle and PE tubing attached to the needle that will be inserted into the appropriate inlets in the chip. This specialized needle allows for gentle flushing and insertion of fluids into the channels.The syringes are then placed into a syringe pump where they can be digitally controlled through a computer software with an accurate flow rate.

In order to optimize flow rate, we flowed blue and orange dyes through different inlets and saw them mix within the microfluidic chip. This allowed us to optimize the flow of liquid through two different channels which later merge into a single channel. Following this, we wanted to flow cell suspensions through our chip. We were hoping to isolate single cells within a droplet, however, we ran out of time and could not fully optimize our system to do so (see video and image below - cells flowing through and we were able to introduce dye to the chips)

The next step would be to isolate single cells. However, we were unable to do this due to time constraints.