Self-Replenishing Vascularized Fouling-Release Surfaces

Inspired by the long-term effectiveness of living antifouling materials, we have developed a method for the self-replenishment of synthetic biofouling-release surfaces. These surfaces are created by either molding or directly embedding 3D vascular systems into polydimethylsiloxane (PDMS) and filling them with a silicone oil to generate a nontoxic oil-infused material. When replenished with silicone oil from an outside source, these materials are capable of self-lubrication and continuous renewal of the interfacial fouling-release layer. Under accelerated lubricant loss conditions, fully infused vascularized samples retained significantly more lubricant than equivalent nonvascularized controls. Tests of lubricant-infused PDMS in static cultures of the infectious bacteria Staphylococcus aureus and Escherichia coli as well as the green microalgae Botryococcus braunii, Chlamydomonas reinhardtii, Dunaliella salina, and Nannochloropsis oculata showed a significant reduction in biofilm adhesion compared to PDMS and glass controls containing no lubricant. Further experiments on vascularized versus nonvascularized samples that had been subjected to accelerated lubricant evaporation conditions for up to 48 h showed significantly less biofilm adherence on the vascularized surfaces. These results demonstrate the ability of an embedded lubricant-filled vascular network to improve the longevity of fouling-release surfaces.Engineering and Applied Science

Infused polymers for cell sheet release

Tissue engineering using whole, intact cell sheets has shown promise in many cell-based therapies. However, current systems for the growth and release of these sheets can be expensive to purchase or difficult to fabricate, hindering their widespread use. Here, we describe a new approach to cell sheet release surfaces based on silicone oil-infused polydimethylsiloxane. By coating the surfaces with a layer of fibronectin (FN), we were able to grow mesenchymal stem cells to densities comparable to those of tissue culture polystyrene controls (TCPS). Simple introduction of oil underneath an edge of the sheet caused it to separate from the substrate. Characterization of sheets post-transfer showed that they retain their FN layer and morphology, remain highly viable, and are able to grow and proliferate normally after transfer. We expect that this method of cell sheet growth and detachment may be useful for low-cost, flexible, and customizable production of cellular layers for tissue engineering

Extracellular Vesicle Capture and microRNA Detection

Cancer is one of the leading causes of death in the United states, and there has been much focus on earlier detection through the discovery of novel, easily accessible biomarkers via liquid biopsies. Extracellular vesicles have shown promise as a noninvasive biomarker for disease diagnosis and monitoring, and have become a treasure trove of information because they have been found to carry proteins, DNA, mRNA and microRNA as well surface markers indicative of the their cell origin. Thus developing methods to profile extracellular vesicles and interrogate the contents of these vesicles is a growing area of research and has the potential to develop into a non-invasive diagnostic platform, a liquid biopsy. The aim of this thesis is to develop a system to capture extracellular vesicles and profile the miRNA patterns present within them. First, we develop various amplification strategies in hydrogel particles for microRNA detection, including a colorimetric detection platform that can be translated to point-of-care settings for a liquid biopsy. We also explored other amplification strategies for increased miRNA detection sensitivity including precipitation-based enzymatic signal amplification and strand displacement amplification. Then we develop methods for extracellular vesicle lysis and miRNA detection using a one-pot lysis and miRNA capture method. Extracellular vesicles were isolated from matched diseased and normal donor serum. Using rolling circle amplification, we performed multiplexed miRNA detection and quantification from serum extracellular vesicles. Calibration curves using rolling circle amplification were used to determine miRNA copy number estimates in agreement with other studies in literature with absolute quantification. Finally, we tune hydrogel particle porosity and use novel functionalization techniques to capture extracellular vesicles based on their surface markers. We explored the use of the thiol-acrylate Michael addition reaction for antibody conjugation and optimized it for extracellular vesicle capture. Using these porous, antibody functionalized hydrogel particles, we captured breast cancer serum and match healthy serum extracellular vesicles using two surface markers, paving the way for multiplexed extracellular vesicle surface marker characterization. Porous hydrogel particles have the potential to considerably enhance the workflow for exosome capture and profiling experiments, through multiplexing, fewer sample preparation requirements, and customizable nature, hence furthering extracellular vesicle research. Incorporating the insights from the MIT Sloan Management program, the commercialization potential and current market landscape for extracellular vesicles was analysed. Extracellular vesicles have shown tremendous potential in the field of therapeutics, diagnostics, and for furthering academic research. The market study shows that the field is growing rapidly, with continued investment from venture capital for new companies, as well as corporate acquisitions by legacy players as they look to enter the field. The work presented in this thesis employs the various benefits of hydrogels for biomolecule detection, namely their biocompatibility, solution-like kinetics, nonfouling nature, and tunable chemistry. We believe that this work can be leveraged to improve upon and develop new technologies for extracellular vesicle capture and analysis, leading to more insights into this promising biomarker, eventually leading to earlier and more accurate diagnosis of disease.Ph.D

A platform for multiplexed colorimetric microRNA detection using shape-encoded hydrogel particles

We report a platform utilizing a reporter enzyme, which produces a chromogenic indigo precipitate that preferentially localizes within a hydrogel microparticle. The 3D network of the hydrogel maintains the rapid target binding kinetics found in solution, while multiplexed target detection is achieved through shape-encoding of the particles. Moreover, the precipitate-laden hydrogels can be imaged with a simple phone camera setup. We used this system to detect microRNA (miRNA) down to 0.22 fmol. We then showed the compatibility of this system with real samples by performing multiplexed miRNA measurements from total RNA from matched colon cancer and normal adjacent tissue.National Institutes of Health (Grant 5R21EB024101-02

Quantitative and Multiplex Detection of Extracellular Vesicle‐Derived MicroRNA via Rolling Circle Amplification within Encoded Hydrogel Microparticles

Extracellular vesicle-derived microRNA (EV-miRNA) represent a promising cancer biomarker for disease diagnosis and monitoring. However, existing techniques to detect EV-miRNA rely on complex, bias-prone strategies, and preprocessing steps, making absolute quantification highly challenging. This work demonstrates the development and application of a method for quantitative and multiplex detection of EV-miRNA, via rolling circle amplification within encoded hydrogel particles. By a one-pot extracellular vesicle lysis and microRNA capture step, the bias and losses associated with standard RNA extraction techniques is avoided. The system offers a large dynamic range (3 orders of magnitude), ease of multiplexing, and a limit of detection down to 2.3 zmol (46 × 10-18 m), demonstrating its utility in clinical applications based on liquid biopsy tests. Furthermore, orthogonal measurements of EV concentrations coupled with the direct, absolute quantification of miRNA in biological samples results in quantitative measurements of miRNA copy numbers per volume sample, and per extracellular vesicle

Power-to-Gas Implementation for a Polygeneration System in Southwestern Ontario

Canada has stockpiles of waste petroleum coke, a high carbon waste product leftover from oil production with little positive market value. A polygeneration process is proposed which implements “power-to-gas” technology, through the use of electrolysis and surplus grid electricity, to use waste petroleum coke and biomass to create a carbon monoxide-rich stream after gasification, which is then converted into a portfolio of value-added products with the addition of hydrogen. A model implementing mixed-integer linear programming integrates power-to-gas technology and AspenPlus simulates the polygeneration process. The downstream production rates are selected using particle swarm optimization. When comparing 100% electrolysis vs. 100% steam reforming as a source of hydrogen production, electrolysis provides a larger net present value due to the carbon pricing introduced in Canada and the cost reduction from removal of the air separation unit by using the oxygen from the electrolysers. The optimal percent of hydrogen produced from electrolysis is about 82% with a hydrogen input of 7600 kg/h. The maximum net present value is $332 M when over 75$203 M when the dimethyl ether is capped at 50% production. The polygeneration plant is an example of green technology used to environmentally process Canada’s petroleum coke