Application of microfluidic emulsion technology to biochemistry, drug delivery and Lab-on-a-Chip programmability

Thesis (S.M.)--Massachusetts Institute of Technology, Dept. of Mechanical Engineering, 2005.Includes bibliographical references (p. 129-138).This research applies microfluidic emulsion technology to three diverse problems; biochemistry, drug delivery and lab-on-a-chip programmability. These subjects represent distinct research programs, but the underlying physics of droplet formation, transport and control at low values of the Reynolds and Capillary numbers in multiphase microfluidics allows them to be considered in parallel and supports the flexibility of this technology. Within these stamp-sized elastomeric polydimethylsiloxane (PDMS) microfluidic devices, pressurized immiscible fluids may be combined at a junction of two or more microchannels, combining crossflow and viscoelastic shear, to generate emulsions. Droplet sizes may be tuned from nanometers to microns in diameter, controlled by device geometry and hydrodynamic flow characteristics. The application of droplets as individual bioreactors for biochemical assays is first explored at the device and external sensor level. The goal of this research is to extend on existing approaches and address challenges of platform scalability. Microchannel design strategies are analyzed then fabricated in order to increase sample incubation periods. Using monodisperse droplet formation within microfluidic devices, techniques are developed for the manufacture of drug loaded biodegradable polymeric particles for controlled release of encapsulated ingredients within biological systems. Coupled with the bulk method of solvent evaporation, microspheres with a tunable range of volumes spanning four orders of magnitude are generated and characterized using this rapid and flexible prototyping technique. Finally, a programmable microfluidic system platform using multiphase flows in soft lithography is developed.(cont.) To demonstrate scalability of this approach, a "general- purpose" microfluidic chip is implemented, where underlying mechanisms for sample manipulation can be integrated to develop more complex systems. This research represents a first step to bring high-level control abstractions to the microfluidic realm, with the aim of enabling a new level of scalability and programmability for lab-on-a-chip experiments.by John Paul Urbanski.S.M

Microfluidic tools for metabolomics

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Mechanical Engineering, 2008.Includes bibliographical references (p. 153-160).A primary challenge in embryology is to understand the factors that govern the development of preimplantation (PI) embryos and how these factors relate to embryo viability in the field of in vitro fertilization (IVF). This is particularly important as clinical policy moving towards single embryo transfer (SET) has gained awareness to manage unprecedented numbers of multiple births, such as twins and triplets, resulting from artificial reproductive techniques. Conditions that correlate with developmental potential of candidate embryos are disputed in the field, however, as the requisite data is difficult to obtain.The metabolic profiles of embryos during in vitro culture have been suggested as a key indicator of developmental potential, and approaches have been clinically implemented to select transfer candidates which make the most efficient use of nutrients. Existing microdroplet analysis techniques are accurate and suitable for non-invasive assessment of single embryos. Unfortunately, the process of determining metabolite levels in nanoliters of culture media through fluorometric assays is low-throughput and requires specialized expertise, hindering widespread clinical use of these methods. The goal of this thesis is to develop microfluidics-based approaches for improving metabolic analysis of PI embryos and mammalian cells. This challenge necessitates two competencies: methods for automating chemical assays and methods for supporting cell cultures, which can be integrated with analysis. Contributions include a standalone platform for determining the metabolite use of single embryos. Profiles may be acquired automatically, which reduces significant technician hours and improves repeatability. Techniques are developed for performing embryo culture in the smallest culture volumes to date in microfabricated environments. Microfluidic approaches have enabled culture that outperforms the current state of art approach based on cell count measurements.(cont.) An integrated system is introduced, merging analysis and culture competencies to perform metabolic profiling of separate cultures of mammalian cells in parallel. Finally, new paradigms in microfluidic design are presented based on the concept of vertically integrated architectures, suitable for overcoming density limitations of microfluidic assays. A scalable analysis platform for refining embryo selection has been long warranted and would enable pursuit of the difficult questions relating metabolism and embryo viability as the clinical movement towards SET continues.by John Paul Urbanski.Ph.D

Abstraction Layers for Scalable Microfluidic Biocomputers (Extended Version)

Microfluidic devices are emerging as an attractive technology for automatically orchestrating the reactions needed in a biological computer. Thousands of microfluidic primitives have already been integrated on a single chip, and recent trends indicate that the hardware complexity is increasing at rates comparable to Moore's Law. As in the case of silicon, it will be critical to develop abstraction layers--such as programming languages and Instruction Set Architectures (ISAs)--that decouple software development from changes in the underlying device technology.Towards this end, this paper presents BioStream, a portable language for describing biology protocols, and the Fluidic ISA, a stable interface for microfluidic chip designers. A novel algorithm translates microfluidic mixing operations from the BioStream layer to the Fluidic ISA. To demonstrate the benefits of these abstraction layers, we build two microfluidic chips that can both execute BioStream code despite significant differences at the device level. We consider this to be an important step towards building scalable biocomputers

Integrated genomic characterization of pancreatic ductal adenocarcinoma

We performed integrated genomic, transcriptomic, and proteomic profiling of 150 pancreatic ductal adenocarcinoma (PDAC) specimens, including samples with characteristic low neoplastic cellularity. Deep whole-exome sequencing revealed recurrent somatic mutations in KRAS, TP53, CDKN2A, SMAD4, RNF43, ARID1A, TGFβR2, GNAS, RREB1, and PBRM1. KRAS wild-type tumors harbored alterations in other oncogenic drivers, including GNAS, BRAF, CTNNB1, and additional RAS pathway genes. A subset of tumors harbored multiple KRAS mutations, with some showing evidence of biallelic mutations. Protein profiling identified a favorable prognosis subset with low epithelial-mesenchymal transition and high MTOR pathway scores. Associations of non-coding RNAs with tumor-specific mRNA subtypes were also identified. Our integrated multi-platform analysis reveals a complex molecular landscape of PDAC and provides a roadmap for precision medicine

The FLUXNET2015 dataset and the ONEFlux processing pipeline for eddy covariance data

The FLUXNET2015 dataset provides ecosystem-scale data on CO2, water, and energy exchange between the biosphere and the atmosphere, and other meteorological and biological measurements, from 212 sites around the globe (over 1500 site-years, up to and including year 2014). These sites, independently managed and operated, voluntarily contributed their data to create global datasets. Data were quality controlled and processed using uniform methods, to improve consistency and intercomparability across sites. The dataset is already being used in a number of applications, including ecophysiology studies, remote sensing studies, and development of ecosystem and Earth system models. FLUXNET2015 includes derived-data products, such as gap-filled time series, ecosystem respiration and photosynthetic uptake estimates, estimation of uncertainties, and metadata about the measurements, presented for the first time in this paper. In addition, 206 of these sites are for the first time distributed under a Creative Commons (CC-BY 4.0) license. This paper details this enhanced dataset and the processing methods, now made available as open-source codes, making the dataset more accessible, transparent, and reproducible.Peer reviewe

Abstraction layers for scalable microfluidic biocomputers

Abstract. Microfluidic devices are emerging as an attractive technology for automatically orchestrating the reactions needed in a biological computer. Thousands of microfluidic primitives have already been integrated on a single chip, and recent trends indicate that the hardware complexity is increasing at rates comparable to Moore’s Law. As in the case of silicon, it will be critical to develop abstraction layers—such as programming languages and Instruction Set Architectures (ISAs)—that decouple software development from changes in the underlying device technology. Towards this end, this paper presents BioStream, a portable language for describing biology protocols, and the Fluidic ISA, a stable interface for microfluidic chip designers. A novel algorithm translates microfluidic mixing operations from the BioStream layer to the Fluidic ISA. To demonstrate the benefits of these abstraction layers, we build two microfluidic chips that can both execute BioStream code despite significant differences at the device level. We consider this to be an important step towards building scalable biological computers.

Digital microfluidics using soft lithography, Lab on a Chip,

Although microfluidic chips have demonstrated basic functionality for single applications, performing varied and complex experiments on a single device is still technically challenging. While many groups have implemented control software to drive the pumps, valves, and electrodes used to manipulate fluids in microfluidic devices, a new level of programmability is needed for end users to orchestrate their own unique experiments on a given device. This paper presents an approach for programmable and scalable control of discrete fluid samples in a polydimethylsiloxane (PDMS) microfluidic system using multiphase flows. An immiscible fluid phase is utilized to separate aqueous samples from one another, and a novel ''microfluidic latch'' is used to precisely align a sample after it has been transported a long distance through the flow channels. To demonstrate the scalability of the approach, this paper introduces a ''generalpurpose'' microfluidic chip containing a rotary mixer and addressable storage cells. The system is general purpose in that all operations on the chip operate in terms of unit-sized aqueous samples; using the underlying mechanisms for sample transport and storage, additional sensors and actuators can be integrated in a scalable manner. A novel high-level software library allows users to specify experiments in terms of variables (i.e., fluids) and operations (i.e., mixes) without the need for detailed knowledge about the underlying device architecture. This research represents a first step to provide a programmable interface to the microfluidic realm, with the aim of enabling a new level of scalability and flexibility for lab-on-a-chip experiments