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TRACK 3 TRACK 3 Technical Program

sample. With an optimized counter-flow rate, the concentration mechanism Technical Presentation. NEMB2016-5973
is stable for more than 25 minutes and capable of concentrating DNA over
1000 times, extending the concentration sensitivity of SML-FSHS over 3 Kerim Kaylan, Viktoriya Ermilova, Ravi Yada, Gregory Underhill,
orders of magnitude to 5 ng/mL (150 fM). Using our integrated method, DNA University of Illinois Urbana-Champaign, Urbana, IL, United States
preconcentration, separation, and sizing can all be accomplished without
the requirement of electrodes or applied electric fields.
Microenvironmental regulation plays a crucial role in stem and progenitor
cell fate/function in a variety of developmental and disease contexts. Reg-
10:40am Nanoelectronic Sensor for Rapid Detection of DNA ulation can occur simultaneously via multiple modes, including cell-cell,
cell-substrate, and cell-soluble factor interactions. Although existing meth-
Technical Presentation. NEMB2016-6072 ods provide powerful means of investigating each of these interactions
individually, there remains a need for high throughput approaches capable
Darius Saadat-Moghaddam, Jong-Hoon Kim, Washington State of deconstructing combinatorial regulation by multiple modes. For instance,
University Vancouver, Vancouver, WA, United States the bipotential differentiation of liver progenitor cells underlies liver devel-
opment and bile duct formation as well as liver regeneration and disease.
The completions of a series of genome projects have dramatically increased This fate specification process is dependent on the action of multiple modes
the capacity of genome information. The increasing availability of genetic of microenvironmental regulation in a spatially-dependent and tempo-
rally-sequenced manner. Prior work has shown that TGF-beta and Notch
information demands more simple and rapid methodology for nucleic acid
testing. To this end, cumbersome procedural steps with long amplification signaling as well as extracellular matrix (ECM) composition play key roles in
time (>>1hour) for polymerase chain reaction (PCR) have been a major chal- both this specification process and associated tissue morphogenesis. Thus,
lenge for rapid identification of genetic information. The ideal goal would elucidation of the complete mechanistic details of this process, including
be the development of a rapid, simple detection methodology without interactions between TGF-beta, Notch, and ECM, requires the combinatorial
capabilities described above.
cumbersome sample preparation steps and/or instrumentation. This paper
addresses the challenge of rapidly identifying genetic information. Unlike
other amplification-based detection methods, a nanoelectronic sensor, sin- Utilizing a series of approaches, including co-cultures and cellular microar-
gle-walled carbon nanotubes (SWCNTs) modified nanostructured needle, rays, we identified distinct contributions of different Notch ligands and ECM
is capable of rapidly concentrating and detecting small amounts of DNA, proteins in liver progenitor fate decisions. We demonstrated a cooperative
influence of JAG1 and TGF-beta-1 on cholangiocytic differentiation. We
due to a concentration and reaction step controlled by an electric field. The
nanoelectronic sensor is fabricated by combination of top-down (for chip further established ECM-specific effects using cellular microarrays on poly-
structure) and bottom-up (for SWCNTs assembly) fabrication approaches; acrylamide hydrogel substrates, known to exhibit well-defined material and
a) fabricate microscale-needle structure and b) immobilize SWNCTs onto mechanical properties. These ECM arrays contained 32 distinct combina-
the structure to enhance the charge transduction through DNA. When the tions of collagen I, collagen III, collagen IV, fibronectin, and laminin. In the
same format, we observed that presentation of exogenous Fc-recombinant
nanoelectronic sensor concentrates target DNA to its surface using an elec-
tric field, target analytes in a sample are circulated by electrokinetic flow. JAG1, DLL1, and DLL4 conjugated with Protein A/G (a means of clustering to
Subsequently, the concentrated targets are further attracted to the tip by a improve retention in the hydrogel and cellular recognition) was sufficient to
dipole moment of polarized targets (dielectrophoresis), transporting targets. induce cholangiocytic differentiation. The extent of differentiation induced
The selectivity is conferred by capillary force through the withdrawal of the by arrayed Notch ligands remained dependent on ECM composition. Last,
by combining these Notch ligand arrays with shRNA-based knockdown of
sensor out of the solution. The capillary force excludes molecules and par-
ticles that are >>80% of the width of the tip. λ DNA, a model analyte, is used Notch ligands in progenitor cells, we systematically examined the effects of
to capture on the sensor surface with applying alternating current (AC) field both cell-extrinsic and cell-intrinsic Notch ligand.
(20V peak-to-peak, 5MHz). To check the sensitivity, various concentration of
λ DNA in TE buffer is tested from 100aM to 1pM with 10-fold increment. For Our results highlight the importance of divergent Notch ligand function and
combinatorial microenvironmental regulation in liver progenitor fate speci-
the electrical detection, DI water is utilized to reduce non-Faradaic current,
and thus increase signal-to-noise ratio. The nanoelectronic sensor dipped fication and additionally serve to highlight a cellular microarray platform ca-
into DI water and an electric detection is performed. Upon presence of DNA pable of interrogating multiple modes of regulation simultaneously. Ongoing
on the sensor surface, the current is decreased. To validate the electrical studies aim to delineate the potential cooperation between gradients of me-
measurement results, the optical measurement is also conducted with flu- chanical forces and Notch ligand-mediated progenitor cell differentiation.
orescence microscope. In conclusion, the sensitivity is 100 aM (3.2pg/mL)
with the assay time of 5 min. Based on the mean value of electrical signal, 9:50am Microscale Modeling of Layered Fibrous Networks Ap-
the nanoneedle shows a potential to directly measure the concentration of plied to Biomaterials for Tissue Engineering
DNA in the sample. The nanoelectornic sensor can detect DNA at the con-
centration of 100 attomolar (aM) in a given sample volume (e.g. 5 µL). Since Technical Presentation. NEMB2016-6140
the nanoelectronic sensor does not require nucleic acid amplification, it is
significantly faster and simpler than PCR approaches.
Michael Sacks, The University of Texas At Austin, Austin, TX, United
States, James Carleton, Greg Rodin, University of Texas at Austin,
Austin, TX, United States
3-7
EXTRACELLULAR MATRIX Layered fibrous networks form the primary microstructure of many natural
and engineered tissues. Networks of collagen, elastin, and other fibrous pro-
teins make up the extracellular matrix (ECM), which is the primary structural
Navarro 9:30am - 11:10am component of cartilage, tendons, blood vessels, heart valves, and other soft
tissues. In this work, we introduce a method for modeling materials that have
Session Organizer: Gregory Underhill, University of Illinois Urba- a layered, nano-fibrous network structures. A primary application is in the
na-Champaign, Urbana, IL, United States field of tissue engineering, where high fidelity models of electrospun scaf-
folds are needed to better understand how the network geometry affects
the mechanical and biological function of the tissues that are grown on the
9:30am Cellular microarrays reveal combinatorial effects of scaffolds. Scaffold geometry has a strong influence on the tissue’s macro-
Notch ligands, TGF-beta, and extracellular matrix on liver pro- scopic mechanical behavior, cell proliferation and attachment, nutrient and
genitor differentiation waste flows, and extracellular matrix (ECM) generation.The random walk al- 41
gorithm used to generate the scaffold geometry mimics the electrospinning