Electroporation is an efficient, low-toxic physical method which is used to deliver impermeant macromolecules such as genes and drugs into cells. Genetic modification of the cell is critical for many cell and gene therapy techniques. Common electroporation protocols can only handle small volumes of cell samples. Also, most of the conventional electroporation methods require expensive and sophisticated electro-pulsation equipment. In our lab, we have developed new electroporation methods conducted in microfluidic devices. In microfluidic-base electroporation, exogenous macromolecules can be delivered into cells continuously. Flow-through electroporation systems can overcome the issue of low sample volume limitation. In addition, in our method, electro-pulsation can be done by using a simple dc power supply, without the need for any extra equipment. Furthermore, our microfluidic chips are completely disposable and cheap to produce. We show that electroporation and electroporation-based gene delivery can be conducted employing tapered asymmetric curving channels. The size variation in the channel's cross-sectional area makes it possible to produce electric pulses of various parameters by using a dc power supply. We successfully delivered Enhanced Green Fluorescent Protein, EGFP, plasmid DNA into Chinese Hamster Ovary, CHO-K1, cells in our microfluidic chips. We show that the particles/cells undergo Dean flow in our asymmetric curving channels. We demonstrate that there are three main regimes for particle motion in our channels. At low flow rates (from 0 to ~75μl/min) cells do not focus and they randomly follow stream lines. However, as flow rate increases (~75 to 500μl/min), cells begin to focus into one line and they follow a single path throughout the micro-channel. When flow rate exceeds ~500μl/min, cells do not follow a single line and demonstrate more complex pattern. We show that the electric parameters affect the transfection efficiency and cell viability. Higher electric field intensity results in higher transfection efficiency. This is also true in the cases with longer electroporation duration time. In our experimental work, we executed flow-through electroporation for various duration times (t = 2 ms, 5 ms, and 7 ms), and at various electric field intensities (from 300 to 2200 V/cm) while we utilized different flow rates as well, i. e. 150 μl/min (focused flow) and 600 μl/min (complex flow). To explore the impact of individual electric pulse length and electric pulse number on electroporation results, we designed control channels with straight narrow sections. Cells experience different hydrodynamic forces in straight channels compared to curving channels. Flow pattern and cell focusing were also studied in control channels as well. Also, electroporation on CHO-K1 cells was successfully conducted in control channels. The hydrodynamic forces under the conditions we used do not appear to show substantial impact on transfection efficiency. / Master of Science

The passive particle separation method of elasto-inertial microfluidics have greatpotential in the field of physics, biology and chemistry. The objective of thisdegree project was to understand particle behavior in curved microchannels fornon-Newtonian fluids. This in order to optimize the separation of 1 µm and 2 µmparticles where the end goal is to create an efficient sample preparation method fordiagnosing sepsis. Fluorescent beads were spiked into PEO solutions of differentconcentrations and used in microfluidic PDMS-glass chips with various radii toexamine the influence of curvature and elasticity as well as the flow rate. Theresult indicated an independence of both curvature and elasticity. Reynoldsnumber and Dean number are dependent on the flow rate which results in atrade-off between a high and low flow rate. A low Reynolds number is not enoughto create Dean vortices that can be used to separate particles while a highReynolds number creates strong Dean vortices that can obstruct the focusing. Later, microfluidic silicon-glass chips were used to separate 1 µm and 2 µm beads.The 2 µm particles were able to focus in two different PEO concentrations whereasthe 1 µm particles did not have time to focus entirely. This makes it possible toseparate 2 µm particles along with some 1 µm particles towards one outlet whileleaving another outlet with only 1 µm particles. This is a promising start butfurther optimization is required before being applied to real bacteria separation. / Den passiva partikelseparationsmetoden elastisk tröghetsmikrofluidik har storapotential inom fysik, biologi och kemi. Målet med examensarbetet var att förståpartiklars förflyttning i krökta mikrokanaler för icke-newtonska vätskor. Dettagjordes för att optimera separering av 1 µm och 2 µm partiklar där slutmålet är attskapa en effektiv provberedningsmetod för att diagnostisera sepsis. Fluorescerandepartiklar tillsatta i PEO-l¨osningar av olika koncentrationer anv¨andes imikrofluidiska PDMS-glas chip med olika radier för att undersöka inverkan avkrökning och elasticitet samt flödeshastigheten. Resultatet indikerade ettoberoende av både krökning och elasticitet. Reynolds nummer och Deans nummerär beroende av flödeshastigheten vilket resulterar i en avvägning mellan en hög ochlåg flödeshastighet. Ett lågt Reynolds nummer är inte tillräckligt för att skapaDean virvlar vilket kan utnyttjas för att separera partiklar medan ett högtReynolds nummer framkallar starka Dean virvlar vilket kan hindra fokuseringen. Sedan användes mikrofluidiska kisel-glas chip för att separera 1 µm and 2 µmpartiklar. 2 µm partiklarna lyckades fokusera i två olika PEO-koncentrationermedan partiklarna av 1 µm inte fokuserade fullt ut. Detta gör det möjligt attseparera 2 µm partiklar tillsammans med ett antal 1 µm partiklar mot ett utloppsamtidigt som ett annat utlopp endast innehåller 1 µm partiklar. Det är enlovande start men ytterligare optimering krävs innan det kan tillämpas på faktiskbakterieseparation.

<p>Particle suspensions in viscoelastic fluids (e.g., polymeric fluids, liquid crystalline solutions, gels) are ubiquitous in industrial processes and in biology. In such fluids, particles often acquire lift forces that push them to preferential streamlines in the flow domain. This lift force depends greatly on the fluid’s rheology, and plays a vital role in many applications such as particle separations in microfluidic devices, particle rinsing on silicon wafers, and particle resuspension in enhanced oil recovery. Previous studies have provided understanding on how fluid rheology affects the motion of spherical particles in simple viscoelastic fluid flows such as shear flows. However, the combined effect of more complex flow profiles and particle shape is still under-explored. The main contribution of this thesis is to: (a) provide understanding on the migration and rotation dynamics of an arbitrary-shaped particle in complex flows of a viscoelastic fluid, and (b) develop guidelines for designing such suspensions for general applications.</p> <p><br></p> <p>In the first part of the thesis, we develop theories based on the second-order fluid (SOF) constitutive model to provide solutions for the polymeric force and torque on an arbitrary-shaped solid particle under a general quadratic flow field. When the first and second normal stress coefficients satisfy  <strong>Ψ</strong>₁  = −2 <strong>Ψ</strong> ₂ (corotational limit), the fluid viscoelasticity modifies only the fluid pressure and we provide exact solutions to the polymer force and torque on the particle. For a general SOF with  <strong>Ψ</strong> ₁ ≠  −2 <strong>Ψ</strong> ₂, fluid viscoelasticity modifies the shear stresses, and we provide a procedure for numerical solutions. General scaling laws are also identified to quantify the polymeric lift based on different particle shapes and orientation. We find that the particle migration speed is directly proportional to the length the particle spans in the shear gradient direction (L_sg), and that polymeric torques lead to unique orientation behavior under flow.</p> <p><br></p> <p>Secondly, we investigate the migration and rotational behavior of prolate and oblate spheroids in various viscoelastic, pressure-driven flows. In a 2-D slit flow, fluid viscoelasticity causes prolate particles to transition to a log-rolling motion where the particles orient perpendicular to the flow-flow gradient plane. This behavior leads to a slower overall migration speed (i.e., lift) of prolate particles towards the flow centerline compared to spherical particles of the same volume. In a circular tube flow, prolate particles align their long axis along the flow direction due to the extra polymer torque generated by the velocity curvature in all radial directions. Again, this effect causes prolate particles to migrate slower to the flow centerline than spheres of the same volume. For oblate particles, we quantify their long-time orientation and find that they migrate slower than spheres of the same volume, but exhibit larger migration speeds than prolate particles. Lastly, we examine the effect of normal stress ratio ? <strong>α</strong>  = <strong>Ψ</strong> ₂ /<strong>Ψ</strong>₁on the particle motion and find that this parameter only quantitatively impacts the particle migration velocity but has negligible effect on the rotational dynamics. We therefore can utilize the exact solution derived under the corotational limit (?<strong>α</strong> = −1/2) for a quick and reasonable prediction on the particle dynamics.</p> <p><br></p> <p>We next experimentally investigate the migration behavior of spheroidal particles in microfluidic systems and draw comparisons to our theoretical predictions. A dilute suspension of prolate/oblate microparticles in a density-matched 8% aqueous polyvinylpyrrolidone (PVP) solution is used as the model suspension system. Using brightfield microscopy, we qualitatively confirm our theoretical predictions for flow Deborah numbers 0 < De < 0.1 – i.e., that spherical particles show faster migration speed than prolate and oblate particles of the same volume in tube flows.</p> <p><br></p> <p>We finally design a holographic imaging method to capture the 3-D position and orientation of dynamic microparticles in microfluidic flow. We adopt in-line holography setup and propose a straightforward hologram reconstruction method to extract the 3-D position and orientation of a non-spherical particle. The method utilizes image moment to locate the particle and localize the detection region. We detect the particle position in the depth direction by quantifying the image sharpness at different depth position, and uses principal component analysis (PCA) to detect the orientation of the particle. For a semi-transparent particle that produces complex diffraction patterns, a mask based on the image moment information can be utilized during the image sharpness process to better resolve the particle position.</p> <p><br></p> <p>In the last part of this thesis, we conclude our work and discuss the future research perspectives. We also comment on the possible application of current work to various fields of research and industrial processes.</p> <p><br></p>