Poster Presentations at AIPP Horizons 3D Bioprinting: Physical and Chemical Processes
Embassy Suites, 460 N Cherry St, Winston-Salem, NC 27101
May 2, 2017
Title: 3D printing of hydrogel scaffolds to support an in vitro model of the human cleft palate.
By: Naveed Saeeed, Paul Cooper, Richard Shelton, Gabriel Landini, Joseph Hardwicke
Affiliation: University of Birmingham
Poster Number: 1
Abstract: INTRODUCTION: Cleft lip and/or palate have an incidence of 1/700 in the UK. The spectrum of
defects range from isolated clefts of the lip or palate to larger clefts involving both, resulting from a
failure of midline fusion in utero. Surgical reconstruction requires lateral palatal tissues to be mobilized
and joined at the midline at the age of six months. Despite improvements in cleft palate surgery,
oronasal fistulae occur as a result of wound breakdown. We anticipate that an autologous cell-based
construct can provide a filler between the oral and nasal mucosal repairs, to reduce fistulation.
METHODS: Primary cleft lip and palatal fibroblasts were isolated from infants undergoing cleft
lip and palate repair at the age of three and six months respectively. Dental impressions taken at the
time of cleft repair were scanned using a Nextengine 3D scanner. Alginate was optimised with regards to
cross-linking to allow for 3D printing of alginate-encapsulated fibroblasts, using a Seraph Robotics
Fab@Home M4 platform.
RESULTS: 18 fibroblast cell lines were cultured from cleft lip (n=12) and palatal (n=6) biopsies.
Alginate was partially cross-linked with the addition of calcium chloride to allow for the even extrusion
of the hydrogel in a manner that would retain a 3D form until completion cross linking. There was no
significant difference between medical and non-medical grade alginate with respect to shrinkage during
the cross linking process. Cells remained viable at 7 days.
DISCUSSION: This proof of concept study has optimised the in vitro culture conditions for
generation of a cell-laden tissue engineered 3D construct and to develop the printing process for the
synthesised tissue. Autologous cells harvested at time of cleft lip repair at three months of age could be
seeded onto a patient specific scaffold to be returned to the patient at cleft palate repair aged six
months.
Title: Craniofacial bone regenerative guided by 3D printed architecture
By: Sang Jin Lee (1), Carlos Kengla (2), James Yoo (2), Anthony Atala (1)
Affiliation: 1) Wake Forest Institute for Regenerative Medicine, 2) Wake Forest School of Medicine
Poster Number: 2
Abstract: Guided bone regeneration strategy utilizes a biomimetic architecture to resist soft tissue
ingrowth while localizing material placed in the site to promote bone growth. In this study, we tested
the effect of guided bone regeneration in a mandibular bone defect of rabbits with a 3D bioprinted
biomimetic architecture paired with an internal architecture designed to facilitate bone ingrowth. Bone
scaffolds were made of a composition of poly(ε-caprolactone) and beta-tricalcium phosphate (PCL/TCP)
and fabricated on the integrated tissue and organ rinting (ITOP) system. We examined bone
regeneration in a critically sized defect at 4 and 8 weeks with a dual architecture bone scaffold and
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inhibited bone regeneration in a defect treated with a scaffold without the soft tissue resistant
architecture. The results showed the increased bone density and volume and new bone formation and
maturation with time. We demonstrated that the printed bone scaffolds were able to organize into
mature tissues of their specific characteristics in vitro and in vivo.
Title: Novel 3D bioprinted model to recapitulate the bone marrow perivascular niche for investigation of
breast cancer dormancy
By: Caitlyn Moore (1), Khadidiatou Guiro (2), Pranela Rameshwar (2)
Affiliation: 1) Rutgers Graduate School of Biomedical Sciences, 2) Rutgers-NJMS Department of
Medicine, Division of Hematology/Oncology
Poster Number: 3
Abstract: Metastatic breast cancer (BC) cells (BCCs) show preference for the bone marrow (BM) which
generally results in poor prognosis. Upon entering the BM, BCCs first interact with cells in the
perivascular niche surrounding the blood vessel and then move toward the endosteal niche adjacent to
the long bone. As BCCs move toward the central region of the BM cavity, they interact with the resident
BM cells directly (e.g. gap junctional intracellular communication) and indirectly (e.g. exosomes and
cytokines), resulting in dormancy. As dormant cells, the BCCs are cycling quiescent and chemoresistant
which gives them a significant survival advantage. This study hypoth
esizes that miRNA-containing exosomes released from perivascular cells are key to BCCs initiating the
transition into dormancy. To recapitulate the in vivo perivascular niche, we developed a hematopoieticsupporting bioink to 3D bioprint BM perivascular niche-mimicking scaffolds. These scaffolds are
biomimetic and able to maintain the major perivascular niche cell populations: endothelial cells (ECs),
mesenchymal stem cells (MSCs), and CXCL12-abundant reticular (CAR) cells. The results showed an
established bioprinted model of perivascular cells that sustains viability and controls cell proliferation.
This study is relevant as it could lead to the identification of targets to prevent dormancy at an early
stage when BCCs enter the BM and during treatment to prevent BCC cycling quiescence.
Title: Design of self-assembling bio-inks for cell-based 3D printing
By: Karen Dubbin, Anthony Tabet, Yuki Hori, Kazuomori Lewis, Sarah Heilshorn
Affiliation: Stanford University
Poster Number: 4
Abstract: Despite the rise of 3D printing of thermoplastics both in industry and the general public, a key
limitation preventing the widespread use of cell-based 3D printing is the lack of suitable bio-inks that are
cell-compatible and have the required properties for printing. Current commonly used biomaterials have
distinct limitations when used as a bio-ink including difficulty maintaining a homogeneous cell
suspension, avoiding cell damage during extrusion, customizing the printed matrix properties to
facilitate cell-matrix interactions, and printing within a bath to prevent cell dehydration while preserving
high print resolution. We designed a new family of tunable biomaterials specifically designed for cellbased 3D printing. These hydrogel-based bio-inks are produced from a blend of engineered recombinant
proteins and peptide-modified, alginate polysaccharides. The use of engineered proteins provides
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control of ligand presentation for cellular attachment and signaling, while the alginate enables cytocompatible, rapid crosslinking within a hydrated sample. This design is advantageous due to the ability
of the bio-ink to undergo two-stages of crosslinking: (i) weak, peptide-based, self-assembly to
homogeneously encapsulate cells within the ink cartridge, to prevent cell settling in the print cartridge,
and to mechanically shield the cells from damaging forces during extrusion and (ii) electrostatic
crosslinking of alginate upon printing within a calcium bath to rapidly stabilize the construct and to tailor
the final mechanical properties for optimal cell-matrix interactions with full cell hydration. Furthermore,
pre and post-print material properties can be varied by altering the alginate or peptide components
individually, leading to a wide range of bio-inks for different tissue engineering applications.
Title: 3D printed acid-cleavable hydrogels using vat photopolymerization
By: Donald Jr Aduba, Evan Margaretta, Alexandra Marnot, Nicholas Chartrain, Katherine Valentine,
Wyatt Surbey, Keyton Feller, Abby Whittington, Timothy Long, Christopher Williams
Affiliation: Virginia Tech
Poster Number: 5
Abstract: Acid-cleavable crosslinking agents comprised of tertiary alkyl esters were incorporated into
Polyethylene Glycol Methacrylate photocurable monomer resins at different weight percent
concentrations to yield degradable covalent networks upon photocuring. These networks were
selectively photo-crosslinked into solid and cellular lattice shapes using a mask-projection microstereolithography (MP-µSLA) process. This study investigates how geometry and acid-cleavable
crosslinker amount dictate their dissolution profile in neutral (pH ~ 7) and acidic media (pH ~ 4), relevant
to healthy and cancerous physiological environments respectively. In addition to looking at dissolution
properties, we explore how crosslinker amount affects material properties such as viscoelasticity, water
sorption, and thermal stability. The biocompatibility of these materials is also evaluated using an MTT
viability assay to determine if these class of 3D-printed networks are suitable for drug delivery
applications. This work presented aims to fundamentally investigate the structure-property-function
relationship of these novel class of 3D-printed biomaterials.
Title: Upregulation of PTEN by dimethyl sulfoxide (DMSO): a good strategy for regenerative medicine
By: On-Yu Hong, Hye-Yeon Jang, Jong-Suk Kim
Affiliation: Chonbuk National University Medical School
Poster Number: 6
Abstract: DMSO is an amphipathic molecule with a highly polar domain and two apolar methyl groups,
making it soluble in both aqueous and organic media. It is one of the most common solvents in
biological studies and medical therapy. The tumor suppressor PTEN (phosphatase and tensin homologue
deleted on chromosome 10, 10q23.3) controls homogeneity of cell junction contractility and hexagonal
cell packing. DMSO has been known to induce upregulation of PTEN, indicating which DMSO play a
pivotal role in the healing of skin wound through up-regulation of PTEN. Here, we demonstrate that
DMSO controls migration of human dermal fibroblast, one of key findings of wound healing model. We
investigated how DMSO regulates cell migration through up-regulation of PTEN in wound model. Up3
regulation of PTEN by DMSO in fibroblasts induces decrease in cell migration. In addition, DMSO caused
reduction of cellular ROS level of fibroblasts. These findings suggest that DMSO might be a promising
material for tissue remodeling in regenerative medicine.
Title: Numerical Investigation of Electrohydrodynamic Bioprinting Process for Cell Deformation
Reduction
By: Ping He
Affiliation: Lamar University
Poster Number: 7
Abstract: A cell-laden droplet impact on to a hydrogel pool with the assistance of electrohydrodynamic
(EHD) forces has been investigated using continuum numerical simulations. Different EHD signals have
been rigorously tested during the printing process, and because of the surface-force nature of both the
EHD and surface tension forces, the EHD method can be used to reduce or even cancel the effect of
surface tension force during the droplet impact process. Our results show that the EHD method is
effective to reduce the cell deformation during the impact process, and thus, it is possible to utilize EHD
to improve the survival rate of dedicate mammalian cells in the violent printing process.
Title: Computational Simulation of Cellular Processes and Oxygenation during Spheroid-Based
Bioprinting
By: Nicanor Moldovan
Affiliation: IUPUI
Poster Number: 8
Abstract: An emerging approach in biofabrication is the creation of 3D tissue constructs through
scaffold-free, cell spheroid-only methods. The basic mechanism in this technology is spheroid fusion,
which is driven by the minimization of energy, the same biophysical mechanism that governs spheroid
formation. However, other factors such as oxygen and metabolite accessibility within spheroids, impact
on spheroids properties and on their ability to form larger-scale structures. The goal of our work was to
develop a simulation platform eventually capable to predict the conditions that minimize metabolismrelated cell loss within spheroids. To describe the behavior and dynamic properties of the cells in
response to their neighbors and to transient nutrient concentration fields, we used an agent-based,
Cellular Potts-type model, applied to a randomly populated spheroid cross-section of prescribed celltype constituency. This model allows a description of: i) cellular adhesiveness and motility; ii)
interactions with concentration fields, including diffusivity and oxygen consumption; and iii)
concentration-dependent, stochastic cell dynamics, driven by metabolite-dependent cell death. Our
model readily captured the basic steps of spheroid-based biofabrication (as specifically dedicated to
scaffold-free bioprinting), including intra-spheroid cell sorting, defect closure, and inter-spheroid fusion.
Moreover, we found that when hypoxia occurring at the core of the spheroid was set to trigger cell
death, this was amplified upon spheroids fusion, but could be mitigated by external oxygen
supplementation. In conclusion, optimization and further development of 3D bio-printing techniques
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could benefit from our computational model able to predict the complex behavior and metabolism of
spheroids-based cellular structures during scaffold-free bioprinting.
Title: A new approach to enhance cellular penetration in 3D hybrid polymer-tissue scaffolds using lasers
By: Jahnavi Sarvepalli, Rama Verma, Bhuvanshwar G S
Affiliation: Indian Institute of Technology Madras
Poster Number: 9
Abstract: Tissue engineering (TE) involves utilization of a wide range of scaffold materials ranging from
polymers, metals, ceramics and hybrid materials for various applications. The properties of these 3D
scaffolds can be tailored according to the application by using a combination of substrates. The use of
biological matrices holds promise for TE but lacks mechanical strength. Hybrid tissue engineered
scaffolds constituting polymeric nanofibers and biological tissues are recently explored that have
attractive bio-mechanical properties. However, they suffer from poor cellular infiltration through small
pores due to dense overlapping nanofibers. We report use of nanosecond pulsed laser to fabricate
micro-scale features on a hybrid scaffold composed of Polycaprolactone-Chitosan nanofiber layered
bovine pericardium to achieve enhanced cellular penetration. The laser energy parameters on the
hybrid scaffolds were optimized to get structured micro-channels with a fluence of 25 J/cm2, 0.1 mm
instep and 15 mark time. Laser irradiation time of 40 μs along with optimized energy parameters
resulted in micro-channel width of ~50 μm and spacing of ~35 μm between adjacent lines. The
mechanical and biological properties of the hybrid scaffolds were not significantly affected after laser
ablation reflecting in the matrix stability. Human mesenchymal stem cells and mouse cardiac fibroblasts
seeded on these laser-ablated hybrid scaffolds exhibited biocompatibility and increased cellular
adhesion in micro-channels. This ultimately led to enhanced cellular penetration into the scaffolds when
compared to non-ablated hybrid TEd scaffolds. These findings suggest the feasibility to selectively ablate
polymer layer in the hybrid TEd scaffolds scaffolds without affecting their bio-mechanical properties and
also describes a new approach to enhance cellular penetration in the 3D hybrid TEd scaffolds scaffolds.
Title: A bladder on a chip with precise deposition for patient’s specific cancer microenvironment
replication
By: Julio Aleman, Cristina Ivan, Anthony Atala
Affiliation: Wake Forest Institute for Regenerative Medicine
Poster Number: 10
Abstract: Bladder cancer has one of the highest urological recurrence rates, due to the lack of noninvasive surveillance methods and low specificity therapies. To optimize current bladder cancer
therapies; we have examined the use of a bladder-on-a-chip as our tissue/disease models. The aim of
our study is to recapitulate a humanized in vitro bladder model in a 3D structure. This platform will be
the foundation towards personalized medicine of bladder cancer modeling, serving to high throughput
patient specific drug screenings.
The chip consists of three layers, the top layer flows bladder smooth muscle specific media, the
middle layer supports the engineered bladder smooth muscle (BSM) layer and urothelial monolayer
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(UM) construct, and the bottom layer flows urothelial specific media modified to resemble human urine.
Healthy bladder organoids were constructed using four different human cell types (bladder urothelial
cells, bladder smooth muscle cells, bladder fibroblasts, and umbilical vein endothelial cells) that were
cultured in an Aggrewell system. Bladder cancer organoids consist on a infusion of T24 cells to healthy
organoids. The UM was seeded on the bottom part of the trans-well membrane by deposition. The BSM
layer was 3D bioprinted in the top part of the membrane, using a personalized hydrogel loaded with
smooth muscle cells. Healthy and cancer organoids were printed into the tissues in the middle layer of
our bladder-on-a chip platform.
We have fabricated a three-layer microfluidic device that replicates the human bladder with the
most concurrent cancer locations (smooth muscle layer, urethelium or interface). Fusion deposition
modeling allows us to generate aligned smooth muscle tissues and to implant an organoid (healthy or
cancer) on any desired layer or position of interest. Overall this platform mimics the bladder’s cancer
microenvironment in a precise, scalable and biologically relevant frame, granting it high throughput
screening competence for patient’s specific therapies.
Title: Highly functionalized and Photoprotective GelMA for 3D BioPrinting
By: Shiny Velayudhan, Anil Kumar Pr, Kalliyana Krishnan V, Kumary Tv
Affiliation: Sree Chitra Tirunal Institute for Medical Sciences and Technology, Trivandrum
Poster Number: 11
Abstract: Gelatin methacrylamide (GelMA) has been increasingly considered as an important bioink
material due to its tailorable mechanical properties, good biocompatibility, ability to photopolymerize in
situ as well as printability. However, a photoinitiated polymerization reaction often leads to radical
accumulation causing dramatic effect upon both the degree of polymerization and post-encapsulation
cell viability. In order to improve the degree of polymerization and the cell viability, we report a multimaterial, cell-compatible bio-ink based on GelMA for 3D bioprinting of customizable tissue and organ
constructs. Our novel bioink system consists of GelMA, UV photoinitiator, 2-hydroxy-1-[4-(2hydroxyethoxy) phenyl]-2-methyl-1-propanone-1-one (Irgacure 2959), along with a cocktail of free
radical scavenger (FRS) molecules. Highly functional GelMA was synthesized using a modified buffer
based technique. Different compositions of bioinks were prepared by mixing GelMA, Irgacure and FRS at
varying ratios. The bioink thus obtained was characterized for its printability, viscosity, gelling
characteristics, mechanical properties and cytocompatibility. The results showed that this novel free
radical scavenging bioink improved relative cell survival while retaining the printability properties.
Title: 3D Printed Hepatic Lobule-Like Bioreactor for in vitro Hepatotoxicity Testing
By: Anil Kumar Pr, Shiny Velayudhan, Roopesh R Pai, Kalliyana Krishnan V, Kumary Tv
Affiliation: Sree Chitra Tirunal Institute for Medical Sciences and Technology
Poster Number: 12
Abstract: In vitro toxicity analysis is one of the most important and simplest methods used to investigate
the potential adverse effects of chemicals and drugs on cells. The extensively used model is the routine
two-dimensional monolayer culture, which results in the loss of cell viability and decreased liver-specific
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functionality. In vitro models such as spheroid culture, sandwich culture and 3D printed tissues have
been proposed to overcome these limitations. A simplified bioreactor by three dimensional (3D) printing
technology specifically for hepatotoxicity has, however, not been reported. In this scenario, we propose
a 3D Printed hepatic lobule-Like bioreactor for in vitro Hepatotoxicity testing. The multi chamber in the
perfusion device can hold functional hepatocytes and is nourished by flow conditions mimicking the in
vivo liver lobule. Simulation of fluid flow by computerized fluid dynamics confirms that the device mimic
the flow conditions of liver lobule. The hepatocytes can be bioprinted using a suitable bioink, within the
chambers, similar to 3D bioprinting inside multiwell plates. The device can perform like a miniaturized
disposable bioreactor for hepatotoxicity testing.
Title: Directing the self-assembly of tumor spheroids within mechanically tunable cellular
heterogeneous in vitro models
By: J. Matt Kinsella, Tao Jiang, Salvador Flores Torres, Jacqueline Kort Mascort, André Charbonneau
Affiliation: McGill University
Poster Number: 13
Abstract: Breast cancer patients with tumors that do not express endocrine receptor, progesterone
receptor or human epidermal growth factor receptor-2 markers represent about 15% of patients and
form the triple negative (TN) subclass associated with poor survival and increased recurrence. We now
understand that tumors are heterogeneous with constant contact of various stromal cell types and that
the tumor microenvironment plays key roles in tumor evolution and resistance to therapy. Here we
report the ability of an extrusion bioprintable composite hydrogel consisting of ionically crosslinkable
alginate and gelatin to promote the formation of multicellular tumor spheroids (MCTS) without the use
of chemical or physical stresses. Multi-cartridge extrusion bioprinting allows us develop 3D cellular
heterogeneous samples comprised of TN breast cancer cells and fibroblasts with specific initial locations
with controlled density. We characterized MCTS progression by quantifying MCTS surface area during a
30-day culture period.
Rheology results indicate a gelling point at 30.6 °C with a period of optimal printing occurring
between 50 to 90 min where the storage modulus is between 96 and 166 Pa and complex viscosity
between 20 and 30 Pa∙s. During this window the hydrogel shows rapid recovery of integrity after shear
in a thixotropic test insuring stability of the substrate. An elastic modulus of ~10 kPa at 37 °C provides
mechanical stability while surface and internal porosity provide a large surface area and greater
exchange rates of essential nutrients and gases. Cancer spheroids form after 7 days of culture with
increasing size over time. After 21 days, fibroblasts migrate to the region where cancer cells were
initially printed and surround the spheroids mimicking stromal symbiotic tumorigenesis. After 30 days
the largest MCTS achieve a surface area of ~80,000 μm2.
Title: 3D Bioprinting of Polymer Bioactive Glass Composite and MSCs for Bone Repair
By: Krishna Kolan, Caroline Murphy, Jakeb Baldridge, Julie Semon, Delbert Day, Ming Leu
Affiliation: Missouri University of Science and Technology
Poster Number: 14
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Abstract: A major limitation of synthetic bone repair is insufficient vascularization of the interior region
of the scaffold. In this study, we investigate the 3D printing of adipose tissue derived mesenchymal stem
cells (AD-MSCs) with polycaprolactone (PCL)/Bioactive glass composite to offer a three-dimensional
environment for complex and dynamic interactions that govern the stem cell’s behavior in vivo. Borate
bioactive (13-93B3) glass was added to a mixture of PCL and chloroform to make an extrudable paste.
Hydrogel mixed with AD-MSCs was extruded as filaments and deposited in between the PCL+glass
filaments. Scaffolds measuring 10x10x1 mm3 in overall dimensions with a filament width of ~400 µm
and pore size of ~300 µm was fabricated. The formation of a hydroxyapatite-like layer on the scaffold
surface was observed after soaking in alpha-Minimum Essential Medium. The scaffold weight loss and
pH change of the media indicated a controlled release of the bioactive glass from the scaffolds.
Live/Dead assay results after 14 days indicated growth and proliferation of MSCs showing the high
potential of the 3D bioprinted PCL/bioactive glass composite scaffolds for bone repair applications.
Title: 3D Bioprinting: Surviving under Pressure.
By: Dianne E Godar
Affiliation: Food and Drug Administration
Poster Number: 15
Abstract: Because 3D bioprinting using microextrusion was reported to yield cells with low viability
(~40%) after pneumatic pressure (40 psi) printing through stainless steel nozzles, or blunt end needles,
with about 150 µm diameters (28 & 30G), we set out to improve the viability by coating the interior of
the nozzles with silicone. For these studies, we used H9 human lymphoma cells to simulate human stem
cells in suspension. To measure cell viability, we used propidium iodide dye exclusion and flow
cytometry to collect data for 10,000 cells. We tried to improve the viability by coating the inside of the
28 & 30G nozzles (1” length) with silicone to protect the cell membranes from being damaged by the
imperfections in the stainless steel nozzle. However, we discovered silicone coating had little effect on
viability because imperfections in the nozzle were not the problem. Instead, the cells being placed in
hypotonic 3% (w/v) alginate prepared in water prior to printing caused significant cell death (~25%) and
considerably more (≥50%) after simulated printing under pressure >200 psi. The situation was resolved
when the alginate was prepared in isotonic solutions of either phosphate buffered saline or complete
culture media (with 1 mM ethylene diamine tetra acetic acid) using pressures over five times what most
printing procedures use. We found the surrogate stem cell model, H9, has good viability (≥85% with 28G
and ≥75% with 30G for 1” length nozzles) after printing under high pressure (220 psi) if the cells are
prepared in isotonic solutions.
Title: Electrochemical Synthesis of Biodegradable/Biocompatible Scaffolds for Orthopedic Applications
By: Mahmoud Saleh (1), M. A. Al-Omair (1), Mohamed M Saleh (2), A. H. Touny (1)
Affiliation: 1) King Faisal University, 2) Wake Forest University
Poster Number: 16
Abstract: Using modified metallic materials as orthopedic tools is an important target for fixation and
reformation of the bones defects and fractures and its mechanical integrity. Problems associated with
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using metals when they are implanted as replacements for hard tissues hinder their full clinical
applications. Biocompatibility is a major challenge confronting metals during their function as
bioimplant. Coating of metal with calcium phosphate (CaPh) is a unique approach to face the above
challenges. Generally, coating the metal surface enhances and accelerates the process of
osseointegration and also increases its corrosion resistant. These biocompatible coatings not only
provide the implant the necessary tribological properties and the desired corrosion resistance, but also
provide them with the desired superior biocompatibility. Electrodeposition (ED) of calcium phosphate
coatings on metal implants is an alternative process that uses aqueous solutions at low temperatures. It
does not affect the structure of the implant and can be applied to complex shapes. Electrodeposition
technique produces superior properties, such as quick and uniform coating. Moreover, it is possible to
control the thickness and chemical composition of the coating. In this work we propose electrochemical
deposition of CaPh on different metal substrates and in different conditions such as bath composition,
pH current density and temperatures. The thus prepared metal/CaPh will be tested for ex situ
biocompatibility and the effects of the different coating conditions on the morphology and structure of
the obtained CaPh will be introduced.
Title: Microstereolithography of Tissue Scaffolds Using a Biodegradable Photocurable Polyester
By: Nicholas Chartrain (1), Maria Vratsanos (2), Dung Han (3), Justin Sirrine (3), Allison Pekkanen (3),
Timothy Long (3), Abby Whittington (3), Christopher Williams (3)
Affiliations: 1) Virginia Polytechnic Institute and State University, 2) Case Western Reserve University, 3)
Virginia Tech
Poster Number: 17
Abstract: Due to its ability to create complex cellular geometries with extremely fine resolution, mask
projection microstereolithography (MPμSL) can be useful for fabricating designed tissue scaffolds and
other biological constructs for use in Tissue Engineering and Regenerative Medicine. However, few
photocurable materials with low cytotoxicity, adequate cell adhesion, and degradability can be
processed with MPμSL. In this work, we present the fabrication of biocompatible and biodegradable
tissue scaffolds with 50 μm feature sizes from a novel polyester using MPμSL. Poly(tri(ethylene
glycol)adipate) dimethacrylate (PTEGA-DMA) was synthesized and evaluated for its printability. The
curing parameters for printing were identified and scaffolds were fabricated. Optical and electron
microscopy were used to determine the achievable feature sizes and accuracy of printed parts using the
polymer in the MPμSL system. MC3T3-E1 mouse preosteoblasts were seeded on PTEGA-DMA films to
assess adhesion and biocompatibility.
Title: 3D bioprinting of hiPSC-based vascular cardiac tissue
By: Vahid Serpooshan, Daniel Hu, James Hu, Sean Wu
Affiliation: Stanford University
Poster Number: 18
Abstract: Recent findings by our group and others have promised great potential of 3D bioprinting
approach to create vascular networks through maintaining a precise spatial control of cells,
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biomaterials, and small molecules in the 3D space. However, the effect of vasculature design
parameters on cardiac cell survival and function in the 3D tissue remains elusive. In this study, we
hypothesized that precise spatial organization of human induced pluripotent stem cell-derived
cardiomyocytes (hiPSC-CMs) and endothelial cells (ECs) within a 3D bioprinted vascular construct can
facilitate the creation of a perfusable and functional 3D cardiac tissue. We 3D printed hiPSC-CMs that
were encapsulated in GelMA bioink and embedded a 3D vascular network. Constructs were
subsequently perfused with ECs utilizing a 3D printed bioreactor system. Our preliminary data
demonstrated EC attachment and formation of endothelium onto the channels via 3D perfusion. We
proved the feasibility of printing viable and functional hiPSC-CMs and ECs at relatively high spatial
resolution (~50 μm) into the 3D space. Moreover, we are currently examining the effect of vasculature
design and fluid flow on hiPSC-CM and EC viability and function in 3D printed myocardium. Together,
these results establish the design principles that can lead to the creation of the first 3D bioprinted,
patient-specific, vascular myocardium. This approach can be broadly applicable to other tissues and
organs. The bioengineered vascular construct can be utilized for other future hypothesis-generating
research projects in both basic science and clinical research applications.
Title: Biofabrication of Tissue-Specific Extracellular Matrix Proteins to Enhance Skeletal Myocyte
Expansion and Differentiation of Muscle Progenitor Cells
By: Yuanyuan Zhang
Affiliation: WFIRM
Poster Number: 19
Abstract: Skeletal muscle progenitor cells (MPCs) are considered a key candidate for cell therapy in the
treatment of skeletal muscle dysfunction due to injury, disease or age. However, it is challenging to
expand sufficient functional skeletal muscle cells in vitro from a small tissue biopsy because skeletal
myocytes often decrease their phenotypic expression in culture conditions. Thus, we sought to develop
a better culture system for expansion and differentiation of MPCs to be used for bioprinting for
myogenesis or cell therapy in vivo to treat urinary incontinence. Porcine-derived muscle extracellular
matrix (ECM) was generated via decellularization methods with distilled water, 0.2 mg/mL DNAse or 5%
fetal bovine serum, with liver and kidney ECM as controls, respectively. Acellular matrices were
homogenized and dissolved. Each ECM solution was combined with a hyaluronic acid-based hydrogel
decorated with heparin (ECM-HA-HP). Cell proliferation and myogenic differentiation capacity of human
MPCs were assessed when cells grew on each ECM-HA-HP substrate. The skeletal muscle ECM-HA-HP
substrate significantly enhanced human MPC proliferation compared to liver or kidney-ECM-HA-HP
substrates. Numbers of myofibers and myotubules significantly increased on muscle ECM-HA-HP
substrate compared to other gel substrates. Numbers of MPCs expressing specific muscle cell markers
(i.e. myosin, desmin, myoD, and myf5) significantly increased when these cells were cultured on muscle
ECM-HA-HP substrate. In conclusion, skeletal muscle ECM-HA-HP as a culture substrate is an optimal
culture microenvironment similar to the in vivo environment. This makes possible the potential use of
skeletal muscle-derived ECM gel in 3D bio-printing to enhance the skeletal muscle repair or cell-based
therapy for skeletal muscle dysfunction.
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Title: High-resolution and high-speed stereolithographic 3D printing for microfluidics and biochip
By: Hiroyuki Yasukochi, Kentaro Soeda, Hiroharu Tamaru, Hirosuke Suzuki, Junji Yumoto
Affiliation: The University of Tokyo
Poster Number: 20
Abstract: We introduce high-resolution and high-speed stereolithographic (SL) 3D printing technology
called RECILS.
One of the advantages of SL is its fabrication speed, but even so it takes about a whole day to
manufacture an object. In addition, printable object’s design is very limited. Almost all 3D printers with
SL need to peel the object from its resin tank each time with every layer. This process takes a long time
and places high stress on the object. Therefore, if the object had fine structures, it can be easily broken.
One of the solutions for this problem is using CLIP method(Carbon3D). However, now, we solved that
problem by controlling the 1-D liquid level of the photocurable resin (eliminating the need for a large
resin storage tank). A glass cylinder and a planar substrate are positioned to make a one-dimensional
gap height of 10 – 40 μm; with the resin fed into the precisely controlled gap. The UV laser light,
modulated according to the object design, scans along the gap through the glass cylinder. After a line of
cured resin had formed, the planar substrate moves to the right and the next line of a cured resin is
formed. When the line-by-line formation of a layer is complete, the substrate moves up by the gap
height, and the next layer is deposited.
This process offers high resolution (10μm) and easy detachment of objects from the
transmission plate without damage. These attributes allow for the fabrication of complex structures
with fine features. In addition to the fine structure, RECILS offers the maximum fabrication size of
90/120/100 cm. In the talk, 3D micro-channel structure and micro-honeycomb structure will be
presented.
Title: 3-D Bioprinting of Innervated Skeletal Muscle Tissue for Functional Recovery
By: Ji Hyun Kim, Young-Joon Seol, In Kap Ko, James J Yoo, Anthony Atala, Sang Jin Lee
Affiliation: Wake Forest Institute for Regenerative Medicine
Poster Number: 21
Abstract: Bioengineered skeletal muscle tissue can be a promising solution to achieve functional
recovery of volumetric muscle injuries. However, the conventional fabrication methods are limited to
building volumetric tissues with functional cellular organization. More importantly, bioengineered
muscle tissues need to be integrated with the host nervous system following implantation, as failure of
innervation results in muscle tissue atrophy. In this study, we utilized 3D bioprinting strategy to fabricate
volumetric skeletal muscle constructs that mimic native skeletal muscle organization. To facilitate longterm tissue survival and accelerate neuromuscular junction (NMJ) formation, human neural stem cells
(hNSCs) were combined with human muscle progenitor cells (hMPCs) in the 3D bioprinted muscle
constructs. Introduction of hNSCs enhanced the cell viability and tissue maturation, which includes
highly aligned myotube formation in vitro. The implanted bioprinted muscle constructs developed highly
oriented myofibers with integration of host vascular and nerve tissues, and muscle mass and muscle
function were increased in a rat tibialis anterior (TA) excisional model. Our results demonstrate that
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creation of innervated volumetric engineered muscle tissue constructs using the 3D bioprinting system
is feasible, and that the muscle construct can contribute to restoration of muscle functions.
Title: Microextrusion based bioprinting apparatus provides valuable method for exploring the 'selforganization' of mammary epithelial branching morphogenesis
By: John Reid, Peter Mollica, Robert Bruno, Patrick Sachs
Affiliation: Old Dominion University
Poster Number: 22
Abstract: Here we provide a detailed account of the design and fabrication of a microextrusion based
bioprinting platform capable of high-throughput, high resolution bioprinting. This investigation details
how to use precision bioprinting techniques to successfully direct the branching morphogenesis of
human mammary epithelial cells cultured in a 3D hydrogel. Printing process parameters were optimized
to provide a robust method for the spatial control of deposited cellular aggregates ranging from 1 to 100
cells. Experimental results provided the minimum number of cells per aggregate to achieve organoid
formation via 'self-organization'. Printing aggregates under these conditions enabled us to accurately
predict the future structures generated by arrays of printed cells.
Title: Uniform Viable 3D Bioprinted Organoids for Medium Throughput Assays
By: Andrea Mazzocchi (1), Shay Soker (2), Aleksander Skardal (1)
Affiliation: 1) Wake Forest Institute for Regenerative Medicine, 2) Wake Forest School of Medicine
Poster Number: 23
Abstract: An increased need for medium throughput reproducible 3D assays has led us to utilize desktop
bioprinting for organoid development across multi-well plates. This method is advantageous as it allows
us to make a 3D human-based tissue analogs to use for drug candidate testing and personalized therapy
optimization. With methods being refined, this technology can translate from immortalized lines to
patient primary tissue. It is vital that models best recapitulate tissue to produce the most relevant
results. Thus, printed organoids must contain the same amount of bioink with a uniform
concentration of cells within an assay to acquire the most accurate results. Here we have shown that we
are able to print replicable organoids of uniform volume and concentration comparable to traditionally
hand pipetted organoids. By creating a uniform bioink we are able to load the syringe and print quickly
and directly into 24-well and 96-well plates. This technology allows for a medium throughput of
organoids unattainable with hand pipetting due to manual speed and bioink properties. Further more,
we have been able to show that viability and drug response are the same between traditional and
printed organoids ensuring that printing can be used in place of hand pipetted organoids. This data
shows that we are able to bioprint lung, pancreatic, and colorectal tissue for drug studies and further
organoid development at medium throughput. Moreover, we are further integrating this technology
with human primary cells, to generate patient-specific models for personalized medicine.
Title: 3D Printing of Fugitive Liquid Metal Inks for Artificial Microvasculature
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By: Dishit Parekh (1), Collin Ladd (1), Lazar Panich (1), Dr. Khalil Moussa (2), Dr. Michael Dickey (1)
Affiliation: 1) North Carolina State University, 2) 3D Systems, Inc.
Poster Number: 24
Abstract: Over the past 30 years, several 3D printing processes have been developed for patterning
polymers and metals. In the fields of biomedical devices and tissue engineering, 3D printing offers
significant advantages due to its ability to rapidly prototype low-volume or one-of-a-kind components
on-demand based on patient-specific needs. A Lab-on-a-Chip (LOC) device is one such mechanism that
integrates multiple laboratory functions including synthesis, transport, sensing and analysis – on a
miniaturized scale – reducing chemical waste and enabling high-throughput screening. The main
challenge in the development of these systems is the design and fabrication of devices as they need to
be functional as well as cost-effective on a small scale. Here, we present a novel yet simple approach to
build 3D microchannels with vasculature at room temperature by direct-writing liquid metal as a
sacrificial template. The printed scaffolds are embedded in a variety of soft (e.g. elastomeric) and rigid
(e.g. thermoset) 3D printable polymers that are bio-compatible. The formation of a surface oxide skin on
the low-viscosity liquid metal stabilizes the shape of the printed metal for planar and out-of-plane
structures. Using an electrochemical reduction method, we can remove the oxide skin – destabilizing the
ink so that it withdraws from the encapsulating material due to capillary forces, resulting in nearly full
recovery of the fugitive ink. Whereas conventional fabrication procedures (e.g., soft lithography)
typically confine microchannels to 2D planes, the geometry of our printed microchannels can be varied
from a simple 2D network to complex 3D multilayered architectures – without using pre-patterned
masks or expensive processing steps. We can also construct “smart” microfluidics by removing select
portions of the liquid metal from the channels, leaving behind 3D metal features that can be interfaced
as antennas, interconnects, or electrodes leading to inexpensive manufacturing of personalized
biosensors among other LOC devices.
Title: Biological properties of 3D-nanocomposites based on scaffolds of carbon nanotubes in protein
matrix
By: Natalia Zhurbina, Aleksandr Polokhin, Alexander Gerasimenko
Affiliation: MIET
Poster Number: 25
Abstract: The technology for production of biocompatible 3D-nanocomposites based on a protein
matrix, impregnated by scaffold of carbon nanotubes (CNTs) is developed. 3D-nanocomposites are
designed to restore osteochondral defects by stimulating the osteoblasts and chondrocytes growth. The
technology includes laser vaporization of water and protein (collagen or albumin) solution of carbon
nanotubes. Electromagnetic radiation promotes the structuring of CNTs and the formation of CNTs
scaffold. Evaporation takes place in layers of 100-500 microns thick. We used the laser device with
optical system based on the IR diode laser. Laser beam moves over the layer of solution using a threecoordinate system. Moreover, laser system is equipped with temperature control system with the
temperature range of 40-100 ºC to prevent the destruction of the proteins structure. We have
fabricated experimental samples of 3D-nanocomposits and examined their biological properties (in vitro
and in vivo). The cells life cycle during cells incubation with the 3D-nanocomposites was studied using
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atomic force microscopy (AFM). Cells were incubated with experimental samples for 3, 24, 48 and 72 h.
The CNTs effect on the morphology of the cells was studied by AFM. There has been a process of cell
adhesion and fixation to the samples, further spreading and breeding. Cells were found in the division
phase after 24 h. 3D-nanocomposite showed no toxicity in contact with living cells, and the ability to
stimulate cell growth. Also, we studied the biochemical and hematological parameters of small
laboratory animal’s (white rats) blood after subcutaneous implantation of samples. Examination of
systems and organs of the body responsible for the metabolism showed no differences between the
experimental and control groups. Using 3D-nanocomposite we repaired cartilage defect of the rabbit
(size 1x0,5 cm) in 70 days. Created 3D-nanocomposite structures can be used to restore the
osteochondral compounds such as subchondral bone and hyaline cartilage.
Title: Physical processes of laser structuring of 3D nanocomposites based on scaffolds of carbon
nanotubes in protein matrix
By: Aleksandr Polokhin, Natalia Zhurbina, Sergey Selishchev, Vitaly Podgaetsky, Levan Ichkitidze,
Alexander Gerasimenko
Affiliation: National Research University of Electronic Technology
Poster Number: 26
Abstract: The work focuses on creation and research of the 3D nanocomposites – synthetic implant
materials which should stimulate cell growth and cytodifferentiation during the tissue forming process.
The 3D nanocomposites were created by the technology of laser vaporization of water and bovine
serum albumin (BSA) solution of multi-walled (MWCNTs) and single-walled (SWCNTs) carbon nanotubes.
Nanotubes were structured to the scaffolds by laser radiation with wavelengths of 810 and 970 nm. The
scaffolds forming was proved with the molecular dynamic modeling technique on example of binding
CNT open end and region of imperfections of another CNT under laser heating from 40 to 100 ºC. The
BSA amino-acid residues (Glu and Asp) interacted with CNTs was determined from the Raman spectra.
The interaction energy of CNT and neutral BSA molecule was 3.7 kJ/mol. Covalent interaction energy of
CNTs atoms and oxygen atoms of amino-acid residues was ~580 kJ/mol. The increasing of SWCNTs
diameter from 1.4-1.6 nm to 30-40 nm was proved with the scanning electronic microscopy (SEM). It can
be associated with the SWCNTs agglomeration to bunch under Van der Waals force and
functionalization of SWCNTs with BSA during the 3D nanocomposites creating process by laser heating.
The nanohardness of 3D nanocomposites was ~370 MPa, the modulus of elasticity was 4.2 GPa, the
strain recovery was ~40 %. The material density was 1.24–1.28 g/cm3. This properties have influence to
mechanical tissue regeneration and rate of 3D nanocomposites biodegradation. The samples based on
SWCNTs had cavities with dimension of 20-30 nm and the samples based on MWCNTs had cavities with
dimension of 35-40 nm. The biological studies have shown good adhesion of living cells - human
embryonic fibroblasts (FEH) – to the 3D nanocomposites surface. The CNTs influence to the 3D
morphology of individual FEHs was studied with SEM and atomic force microscopy.
Title: A 3D Temporal Bone Template for Composite Graft Bioprinting
By: Austin Rose (1), Ola Harrysson (2)
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Affiliation: 1) University of North Carolina School of Medicine, 2) North Carolina State University, Dept.
of Industrial & Systems Engineering
Poster Number: 27
Abstract: High resolution, 3D-printed temporal bone models have been developed for both surgical
training and pre-operative simulation of challenging surgical cases. We review here the materials and
methods used in the creation of these models, including the use of micro-CT data for improved
resolution, the creation of both adult and pediatric models and the use of patient-specific models for
pre-operative surgical simulation. We propose that these 3D models may also serve as templates for the
future printing of bio-compatible implants and viable, multi-tissue composite grafts for use in temporal
bone reconstruction.
Title: Design of continuous-layered hydrogel scaffolds for pneumatic extrusion-based bioprinting
By: Taneka Jones
Affiliation: University of Illinois Chicago
Poster Number: 28
Abstract: The fabrication of biomimetic soft tissues using pneumatic extrusion based bioprinting (EBB) is
challenged by the limited availability of printable hydrogel bioink materials. Additional additive
manufacturing constraints include repetitive design patterns that do not require filament intersection or
excessive bioink deposition start and stop points. To address these limitations, we designed repetitive
fractal space-filling curve patterns for 3D bioprinted human dental pulp applications using mechanically
tunable bioink components. Photopolymerized porous polyethylene glycol diacrylate-gelatin- hyaluronic
acid (PEGDA-Gn- HA) scaffolds were designed to overcome cellular nutrient diffusion limits of ~200 um.
Here, we varied the design architecture to control scaffold porosity and achieve mechanical support for
subsequent layering. Our results demonstrate that current limitations in nozzle diameter to achieve
desired porosity and continuous layering using EBB may be overcome using fractal space-filling curve
models.
Title: Developing of Human 3D Blood Vessel Organoids
By: Nadia El Akabawy (1,2), Miemie Wan (1), Anthony Atala (1), John Jackson (1)
Affiliation: 1) WFIRM, 2) Zagazig University, Faculty of Medicine
Poster Number: 29
Abstract: Introduction: The blood vessels consist of arteries, arterioles, capillaries, venules, and veins.
The goal of this study is to optimize a protocol for formation a 3D model of blood vessel sheroids to
enable studying different drugs metabolic effects and proteomics.
Materials and Methods: Human aortic smooth muscle cells, human aortic endothelial cells and
human dermal fibroblast were cultured using 96 well plates in 200 ul smooth muscle cell specific media
in a ratio of 6: 3: 1 respectively. Spheroids were made using 1200 cells and reached 200 microns in
diameter. Spheroids were allowed to grow in culture for 4 weeks with weekly evaluation through; Live
and dead staining for confocal evaluation, Paraffin embedding for histology examination and ATP assay.
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Results: ATP assay was maximum at day o then showed fluctuations through 1st, 2nd and 3rd
weeks to reach an elevated level at week 4. Results coincide with confocal microscopy for live and dead
staining. Histological examination of paraffin embed blocks and slides stained with H& E, Masson
trichrome and immunofluorescent staining showed special arrangement of the 3 cell types with
secretory activity of some collagen.
Title: Selective laser melting 3D-printed metallic implants for enhancing Osseointegration
By: Shaomian Yao (1), Mary Bollman (1), Raphael Malbrue (1), Chunhong Li (1), Hong Yao (1), Shengmin
Guo (1), Thomas Lallier (2)
Affiliation: 1) Louisiana State University, 2) Louisiana State University Health Science Center
Poster Number: 30
Abstract: Rationale: Establishment of strong osseointegration is critical for successful bone and dental
implants. Slow osseointegration, incomplete osseointegration and loss of osseointegration over time are
the major problems in dental and bone implant surgeries. Selective laser melting (SLM) 3D-printing is an
additive manufacturing technology that can be used to produce patient-specific metallic implants. We
fabricated implants designed to have an interconnected 3D-structure (I3DS) using SLM based 3Dprinting, and tested the hypothesis that implants with such interconnecting structure can reserve
substances, such as stromal-cell derived factor-1 alpha (SDF-1α) for its sustained release to recruit
endogenous stem cells to the implants to promote osseointegration.
Methods: Implants with pore sizes ranging from 0 to 300µm were coated with SDF-1α. The
implants were subjected to an SDF-1α releasing test by incubating them in medium for designated
periods. The SDF-1α released to the medium was determined by enzyme-linked immunosorbent assay
(ELISA). To determine osseointegration, implants were surgically inserted into the skulls of rabbits. The
animals were sacrificed about 2 to 3 months after surgery, and osseointegration of the implants was
analyzed by reverse-torque test, Alizarin red staining and histology analysis.
Results: The implants with 200 and 300µm interconnecting-pores released SDF-1α more slowly
than the non-porous implants. The I3DS implants coated with SDF-1α resulted in enhanced
osseointegration as compared to the solid implant control.
Conclusions: SLM can be used to fabricate implants with interconnecting 3D porous structures
and such porous structures help to retain SDF-1α leading to enhancement of osseointegration.
Title: Nanoengineered Ionic-Covalent Entanglement (NICE) Bioinks for 3D Bioprinting
By: David Chimene (1), Lauren Cross (1), Charles Peak (1), James Carrow (1), Eli Mondragon (1), Guinea
Cardoso (2), Roland Kaunas (1), Akhilesh Gaharwar (1)
Affiliation: 1) Texas A&M University, 2) University of Campinas
Poster Number: 31
Abstract: A number of advanced hydrogel compositions have been recently developed, including
nanocomposites and ionic-covalent entanglement (ICE) networks, that result in excellent printing
characteristics without compromising cell viability. We have developed a novel Nanoengineered IonicCovalent Entanglement (NICE) bioink that combines these approaches to result in a cytocompatible
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bioink with superior mechanical properties to nanocomposite or ICE bioinks. Specifically, nanosilicates
(nSi) were combined with an ICE network formed from gelatin methacrylate and κ-carrageenan
(GelMA/kCA). Rheological testing of non-crosslinked NICE and the component bioinks (nSi-GelMA, nSikCA and GelMA/kCA) revealed that NICE had the highest yield point of ~200 Pa, indicating that NICE
flows during extrusion, but rapidly solidifies upon printing to retain high print fidelity. UV exposure
solidified all GelMA-containing bioinks within 80 seconds, indicating that the inclusion of kCA and
Laponite nanoparticles did not adversely affect crosslinking kinetics. Lyophilized photocrosslinked NICE
hydrogels reveal a high level of interconnected porosity comparable to that of the component
hydrogels. Accelerated degradation studies showed NICE hydrogels were substantially more stable than
any of these component hydrogels. The addition of nSi to GelMA, kCA and GelMA/kCA hydrogels
significantly increased compressive moduli, suggesting that nanosilicates interact with both GelMA and
kCA polymers to mechanically reinforce the polymer networks. In particular, the NICE hydrogel exhibit a
higher compressive modulus than any of the component hydrogels. A self-supporting cylinder printed
with 100 layers exhibited <10% variability in wall thickness along the cylinder length, illustrating both
the fidelity and mechanical resilience of the crosslinked NICE bioink. NIH 3T3 fibroblasts encapsulated in
the NICE bioink filament structures during printing remained viable for up to 60 days in culture. These
results illustrate that nanosilicates interact with both components of GelMA/kCA ICE networks to result
in cytocompatible NICE bioink capable of printing freestanding structures.
Title: Optimizing the Tissue Engineering of Tubular Organ Structures by Bio-Printing
By: Ashkan Shafiee (1), Gabor Forgacs (2)
Affiliation: 1) Wake Forest Institute for Regenerative Medicine, 2) University of Missouri
Poster Number: 32
Abstract: Optimizing the conditions for cells to self-assemble into functional structures in vitro may help
to engineer tissues and eventually organoids [1,2]. To facilitate self-assembly one can employ the
technology of bioprinting, which is a robust and accurate way to arrange cells and supporting biological
materials [3]. Bioprinting is distinct from 3D printing in that post-printing maturation comprises a critical
part of the tissue or organoid formation. After printing, it takes time for the structure to mature via selforganization processes, in particular tissue fusion, and become fully functional. Quantitative information
about the maturation process is critical to assure that the final product has the appropriate properties
for use [4]. In particular the possibility to accelerate this process may allow for earlier implantation.
Tubular organ structures form a large portion of the human body and play vital role in it. Here we report
on the optimization of the tissue engineering process of fabricating tubular bio-printed biological
structures, which, among others, allows for predictive quantification of the maturation process. We
prepared convenient bioink particles, spherical and cylindrical aggregates and assessed their
biomechanical properties such as apparent tissue surface tension (ATST) and characteristic fusion time.
The fastest fusion of bioink particles and, consequently construct maturation, was achieved using
cylindrical bioink. This finding has important implications for the time needed to fabricate vascular
conduits for regenerative purposes.
[1] Shafiee A, McCune M, Forgacs G, et.al., 2015 Post-deposition bioink self-assembly: a quantitative
study Biofabrication 7 045005
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[2] Shafiee A and Atala A 2017 Tissue Engineering: Toward a New Era of Medicine Annual Review of
Medicine 68, 29-40
[3] Shafiee A and Atala A 2016 Printing Technologies forMedical Applications Trends in Molecular
Medicine 22 254–65
[4] McCune M, Shafiee A, Forgacs G, et. al., 2014 Predictive modeling of post bioprinting structure
formation Soft Matter 10 1790–800
Title: Inkjet Printing of Biocompatible Electronic Based Sensors for Real-Time Monitoring of Tissues
By: Ashkan Shafiee, Anthony Atala
Affiliation: Wake Forest Institute for Regenerative Medicine
Poster Number: 33
Abstract: A thorough comprehension of molecular and electrical mechanisms of interaction among cells
within a tissue is essential to control different biological phenomena. Cost-effective, non-destructive,
and real-time monitoring systems may provide insights at the cellular level, although current
technologies are limited in terms of size and accuracy [1]. Notably, however, cells have been shown to
interact with surrounding electric fields; this property has been harnessed by scientists to influence their
behavior. Various studies have shown that electrical forces can be employed for microscale cell
manipulation [2]. Furthermore, crucial roles for electric fields in cellular- [3], tissue- [4], and organ-based
interventions [5] have been identified. Herein, we report the fabrication of biocompatible, electronic
based sensors that allow real-time monitoring of tissues. We employed biocompatible electronic devices
that can detect electrochemical changes in satellite muscle cells and interpret cellular signals. The speed
and accuracy of such electronic devices enable us to create a platform for real-time monitoring of cell
interactions with high precision.
[1] Tian B, et.al., 2012 Macroporous nanowire nanoelectronic scaffolds for synthetic tissues. Nat Mater
11 986–94
[2] Voldman J 2006 Electrical forces for microscale cell manipulation. Annu. Rev. Biomed. Eng. 8 425–54
[3] McCaig C D, et. al., 2009 Electrical dimensions in cell science Journal of Cell Science 122 4267–76
[4] Zhao M 2009 Electrical fields in wound healing—An overriding signal that directs cell migration
Seminars in Cell & Developmental Biology 20 674–82
[5] Bouton C E, et.al., 2016 Restoring cortical control of functional movement in a human with
quadriplegia. Nature 533 247–50
Title: Effect of Bioactives on Extrusion Based 3D Printing of Soft Degradable Polyesters
By: Tanmay Jain (1, 2), David Saylor (2), Qianhui Liu (1), Viraj Patel (3), Rahul Kaushal (4), Jae-Won Choi
(5), Abraham Joy (6), Irada Isayeva (2)
Affiliation: 1) University of Akron, Polymer Science Department, 2) U.S Food and Drug Administration,
OSEL-CDRH-DBCMS, 3) University of Marlyand, Baltimore, 4) University of Marlyand, College Park, 5)
University of Akron, Mechanical Engineering Department, 6) University of Akron, Polymer Science
Department
Poster Number: 34
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Abstract: 3D printing has enabled bench-top fabrication of customized constructs with intricate
architectures. However, the printing of most biomaterials requires the addition of small molecules like
photo-initiators, rheology modifiers, solvents, as well as high temperatures to achieve optimal print
quality. Such processing conditions raise regulatory questions related to material stability and device
biocompatibility. These questions are amplified when it comes to processing of bioactive materials, i.e.,
materials containing bioactive molecules, such as pharmaceuticals and biologics designed to elicit
desired biological response when implanted.
We recently reported the synthesis and 3D printing of biodegradable soft polyesters without solvent,
monomer, initiator, and at room temperature.1 The ability to print these polymers under benign
conditions made these materials especially attractive for printing of bioactive polymer scaffolds. Though
addition of bioactive fillers may cause the desired biological response, these fillers can significantly alter
the physical properties of a baseline polymer and consequently, the printing process.
In this study we examine the effect of adding a pharmaceutical, dexamethasone, on extrusion based 3D
printing of low modulus degradable polyesters. Specifically, we investigate the effects of drug-polymer
interaction and drug loading on polymer flow behavior, material printability and select performancerelated scaffold properties in order to better understand printing of combination products. We, 1)
characterize drug-polymer interaction via binary phase diagrams constructed by combining the
experimental melting point depression data with Flory-Huggins theory; 2) measure viscoelastic
properties of drug/polymer formulations and examine the relationship between these properties and 3D
printing processing parameters, such as the nozzle-to-substrate gap and the layer height; and finally, 3)
show the effects of drug loading on the degradation and the drug release rate of the 3D printed
scaffolds. References: 1. Macromolecules, 2016, 49 (7), pp 2429-2437.
Title: In Vitro Evaluations of 3-D Bioprinted Cardiac Tissue Constructs
By: Zhan Wang (1), Heng-Jie Cheng (2), Young-Joon Seol (1), James Yoo (1), Anthony Atala (1), Sang Jin
Lee (1)
Affiliation: 1) WFIRM, 2) WFBM Cardiology
Poster Number: 35
Abstract: Bioengineering of functional cardiac tissue constructs composed of cardiomyocytes has many
utilities, including surgical repair and enhancement of cardiac tissue as well as developing an in vitro
tissue model for drug discovery, screening and toxicity studies. However, the complexity of myocardium,
structurally and functionally, still presents many challenges for these applications. The cardiac tissue
possesses highly organized structures with unique physiological and biomechanical properties. In this
study, we applied 3D bioprinting strategy to fabricate functional and contractile cardiac tissue
constructs. Rat neonatal heart tissues were obtained to isolate cardiomyocytes, and the cells were
suspended in a fibrin-based hydrogel bioink. Cell-laden hydrogel was printed through a 300-micron
nozzle by pneumatic pressure. The bioprinted cardiac tissue constructs showed spontaneous contraction
after 3 days post-printing and demonstrated synchronized contraction after 14 days in culture,
indicating of cardiac tissue development and maturation. Cardiac tissue formation was confirmed by
immunostaining with antibodies specific to α-actinin and connexin 43, which showed aligned, dense
matured cardiomyocytes. The bioprinted cardiac tissue constructs also showed physiological responses
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(beating frequency and contraction forces) to known cardiac drugs (epinephrine and carbachol).
Moreover, tissue development of the printed cardiac tissue could accelerate by Notch signal blockade.
Our results demonstrated the feasibility of printing functional cardiac tissues that could be used as a
reliable and reproducible model pharmacological applications.
Title: Diels-Alder Reversible Thermoset 3D Printing: Isotropic Thermoset Polymers via Fused Filament
Fabrication
By: Kejia Yang, Walter Voit
Affiliation: University of Texas at Dallas
Poster Number: 36
Abstract: FFF is one of the most popular techniques in 3D printing due to its relatively low cost and high
prototyping speed, but the quality of the printed parts usually lacks the isotropic toughness and
reliability of injection or compression molded parts due to the poor interlayer adhesion between printed
filaments. FFF printing also requires melt-processable thermoplastic materials, which often leads to poor
mechanical properties in the printed parts. Crosslinked polymers, also called thermosets, generally have
better thermal stability, chemical resistance and mechanical properties than thermoplastics. These
qualities make thermosets desirable for many engineering applications, and will efficiently improve the
interlayer adhesion if crosslinks can be introduced between layers of 3D printed parts. Dynamic covalent
chemistry, specifically the furan-maleimide Diels-Alder (fmDA) chemistry, has been applied to many
types of smart materials, such as self-healing materials, recyclable thermosets and others. It also has
great potentials in the 3D printing applications, due to its reversibility making thermoset polymers meltprocessable in a mild heating environment.
In this presentation, we will be discussing our work on developing a new printing technology based on
the FFF printing, Diels-Alder Reversible Thermoset (DART) 3D printing, dedicated for reversibly
crosslinked polymers, which show smoother surface finish, low anisotropy and reasonable toughness in
the 3D printed parts while maintaining a great melt-processability.
Title: Development of cGMP bioprinting methods for human clinical trials
By: Darren Hickerson, Sarah Albertson, Heather Herron, Todd Meinecke, Almudena Martinez-Fernandez,
Rich Payne, Julie Allickson
Affiliation: Wake Forest Institute for Regenerative Medicine
Poster Number: 37
Abstract: Background: A major goal of 3D bioprinting is production of finished constructs for
implantation in regenerative medicine therapies to meet unmet medical needs. If that is to occur, cGMP
methods must be employed in the bioprinting process.
Methods: Risk analysis was performed to identify areas of risk to human subjects and to assess
cGMP regulatory compliance requirements for 3D bioprinting. In response to the risk analysis, the
following engineering, documentation, or procedural changes were made and put into use for preclinical definitive studies: (1) sufficiency of the existing WFIRM bioprinters was assessed for cGMPcompliant printing relative to other commercially available models; (2) an ISO5 environment was
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created by designing and building a softwall HEPA filtered enclosure; (3) the construct was protected
from particulate shedding by mechanical components of the bioprinter using a surgical drape and
cleaning of bioprinter surfaces before each operation; (4) the cooling system fan was diverted to reduce
particulate dispersion; (5) bioprinting syringes and nozzles were assessed for biocompatibility; (6)
sterilization methods were assessed (pending future validation); (7) documentation support for the
bioprinters was collected and formalized in an equipment folder; (8) formalized, approved equipment
operating procedures and cGMP-compliant batch records were implemented for documentation of
construct production; and (9) biocompatibility of bioink polymer and cell suspension reagents and
components were assessed.
Results: The cGMP-compliant engineering solutions, procedural changes, and documentation
were implemented. The bioprinting environment was tested and determined to meet ISO5 standards.
GLP pre-clinical definitive studies were successfully performed. Some elements identified are still in
development pending qualification for use in human clinical trials.
Conclusions: For human clinical trials, 3D bioprinting must meet cGMP standards. Our risk
analysis, engineering controls, documentation support, materials management, and procedural
solutions were successful in GLP animal studies, and are pending finalization and qualification for human
clinical trials.
Title: Protein containing bio ink for 3D printing photo electrochemical cells.
By: Anu Stella Mathews, Carlo Montemagno, Sinoj Abraham
Affiliation: University of Alberta
Poster Number: 38
Abstract: Our work focus on the development of light curable bioinks by incorporating biological
function as an intrinsic property in the devices we print. This approach allows the 3D printing of acrylic
polymer compositions containing biological materials, especially membrane proteins using a photo
(407nm) curing SLA/DLP 3D printer. Retaining the bio functionality of these proteins gives the fourth
dimension (4D) aspect to this construct. Herein, we report the 4D printing of a bio-inspired nano hybrid
electrode for water-splitting applications using a polymeric resin with proton-pumping protein,
bacteriorhodopsin (bR), silver nanoparticles (Ag NP), and carbon nanotubes (CNT). These printed
photoelectrochemical cells exhibit high durability, low onset over-potential, and upon light irradiation
(545nm) produces hydrogen by a synergistic effect of Ag NP and bR.
Title: Optimization of Hydrogel-based Bioink Printability using Rheological Parameters: A Systematic
Approach
By: Gregory Gillispie, Teng Gao, Joshua Copus, James Yoo, Anthony Atala, Sang Jin Lee
Affiliation: Wake Forest Institute for Regenerative Medicine
Poster Number: 39
Abstract: The dearth of suitable bioinks is frequently cited as a major limiting factor for the
advancement of bioprinting technology. It is well known that the viscosity of a bioink has a direct impact
on its printability. Highly viscous materials maintain their structure better, but also have decreased cell
21
viability due to the high forces which are required for extrusion. However, complex viscosity is
comprised of two discrete components, storage modulus (G’) and loss modulus (G”) and their individual
contributions to printability are unknown. For this study, gelatin and alginate were mixed at various
concentrations to obtain hydrogels with a wide range of storage and loss moduli. These hydrogels were
then evaluated for the quantitatively defined values of extrudability, structural integrity, and extrusion
uniformity. While neither loss nor storage moduli alone were excellent predictors of printability, a lower
loss tangent (G”/G’) typically correlated with increased structural integrity while a higher loss tangent
correlated with increased extrusion uniformity. Hydrogels with a loss tangent in the range of 0.3 to 0.65
exhibited an excellent compromise between structural integrity and extrusion uniformity. For
extrudability, increasing either the loss or storage modulus increased the pressure required to extrude
the bioink. A mathematical model relating the G’ and G” to the required extrusion pressure was derived
based on our data. Using this approach, a variety of different bioink formulations can be quickly and
accurately evaluated for printability. Immediate future work will incorporate cell viability studies to
further define printability and will also examine additional hydrogels to determine the generalizability
our evaluation methodology for the printability of bioinks.
Title: Non-pneumatic Actuation of Stretchable Membranes for Tissue Engineering Applications
By: Kiersten Haffey (1), Tony Huang (2), Shuprio Ghosh (1), Arefin Ayesha (1), Jennifer Harris (1), Pulak
Nath (1)
Affiliation: 1) Los Alamos National Laboratory, 2) National Tsing Hua University
Poster Number: 40
Abstract: This poster presents a microfluidic actuation method to simulate and monitor cyclic
mechanical stretching for tissue engineering applications which can also be applied seamlessly with 3D
printed substrates. The ability to simulate and monitor biological forces such as cyclic stretching is an
important parameter in many physiological models. Such mechanical stretching has been demonstrated
to enhance the recapitulation of 3D tissues including cardiac, muscle, vasculature, ligament, tendon, and
lung. Typical approach for mechanical stretching involves additional hardware such as the application of
pneumatic sources, clamps and holders, magnets, or piezo electric elements. In this work, we present a
novel microfluidic method to actuate stretchable membranes that can be simultaneously used for
perfusion, without additional hardware.
The microfluidic stretching platform has 2 channels stacked on top of each other separated by a
thin membrane. The bottom channel has an inlet and outlet where the cross-section of the outlet is
much larger than the inlet. Pulling media from the inlet to the outlet can produce a pressure drop inside
the bottom channel. This pressure drop causes the membrane to bulge. By pausing the flow in a cyclic
fashion and letting the stretched membrane relax, a cyclic stretch can be applied to the substrate. The
media used to create the pressure drop is also used to perfuse and replenish the embedded cells.
By modulating the flow in the bottom channel, we have demonstrated that the membrane can
be stretched cyclically for extended period- resulting in a versatile non-pneumatic stretching technique
for organs-on-a-chip and 3D printed tissues. The simplicity of this approach also allows multiplexing for
high throughput testing.
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This work was partially funded by the Science Undergraduate Laboratory Internships (SULI)
program by the Department of Energy and the RAM-ATHENA program by the Defense Threat Reduction
Agency (DTRA).
Title: Role of quantum dots on luminescent emission and dielectric behaviour with conducting polymer
By: Chetna Tyagi, Ambika Devi
Affiliation: The North Cap University
Poster Number: 41
Abstract: The luminescent and dielectric performance of cadmium selenide (CdSe) quantum dots with
polymers by making composite was investigated experimentally and theoretically. The water soluble
polymer polyvinyl pyrrolidone (PVP) has been chemically synthesized to make a composite material with
CdSe quantum dots at room temperature. Photoluminescence analysis checks the possible interaction
between CdSe quantum dots and PVP matrix. Red shift of photoluminescence spectra of polymer
nanocomposite for CdSe nanoparticles in comparison to bulk CdSe. Dielectric constant (ε') value as the
temperature is increased which corresponds to polar dielectrics. Orientation of dipoles themselves with
increased temperature leads to increased dielectric constant. Dielectric losses is calculated which relates
to moment of charge carriers and dielectric relaxation of material. Dielectric constant frequency
variation is studied by Cole-Cole theoretical model. The value of m, τ, β, σsc is obtained by fitted graphs
which resembles the experimental data
Title: Scaffolds in tissue engineering articular cartilage
By: Laila Montaser (1), Hadeer Abbassy (2), Sherin Fawzy (3)
Affiliation: 1) Menoufia University-Egypt, 2) Alexandria University, 3) Stem Cell, Regenerative Medicine,
Nanotechnology and Tissue Engineering Group (SRNT)
Poster Number: 42
Abstract: Articular cartilage possesses only a weak capacity for repair; therefore, regeneration of its
defects is considered one of the most important challenges. Articular cartilage generation by
autogenous cell/tissue transplantation is one of the most promising techniques in orthopedic surgery
and biomedical engineering. Treatment concepts based on those techniques would eliminate problems
of donor site scarcity, immune rejection and pathogen transfer. Mesenchymal stem cells (MSCs)
obtained from the patients can be expanded in culture and seeded onto a scaffold that will slowly
degrade and resorb as the tissue structures grow in vitro and/or in vivo. MSCs are specified as
appropriate cell candidates for regenerating incurable defects of articular cartilage. The scaffold
provides the necessary support for cells to proliferate and maintain their differentiated function, and its
architecture defines the ultimate shape of the new cartilage. Conventional fabrication methods used for
manufacturing three-dimensional (3D) scaffolds, such as electrospinning, fiber deposition, freeze-drying,
gas foaming, and salt leaching, lack precise control of internal structural features and topology.
Therefore, techniques for the accurate fabrication of multifunctional scaffolds are needed. Considering
the inherent shortcomings of conventional scaffold-based tissue repair, a new bio-fabrication approach,
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termed “(3D) bioprinting,” has been introduced in regenerative medicine. The role of bioprinting is
emerging as an innovative technology that allows for the creation of organized 3D tissue constructs.
Bioprinting technology shows potential in tissue engineering for the fabrication of scaffolds, cells, tissues
and organs reproducibly and with high accuracy. This process also has the capability to combine cells
and biomaterials in an ordered and predetermined way. In this paper, emergent approach in the field of
articular cartilage tissue engineering is presented for specific application. In the next years, the
development of new strategies using stem cells, in scaffolds, with supplementation of culture medium
could improve the quality of new formed cartilage.
Title: Polymer Scaffold for Artificial Trachea
By: Lucero Ramirez, Megan Li, Kejia Yang, Walter Voit
Affiliation: University of Texas at Dallas
Poster Number: 43
Abstract: Polymer scaffolds have emerged as a potent tool in various areas of tissue engineering. Recent
advances have shown that they can be employed as versatile 3-dimensional matrixes for the cultivation
of cells and in the formation of targeted tissue growth. Hydrogels, in particular, show immense potential
as scaffold materials based on their structural similarity to the extracellular matrix of several tissues. In
this project, a biocompatible block copolymer is synthesized based on thiol-ene reaction. The thiol-ene
reaction is also described as a thiol-click chemistry: conditions that identify high-yield and facile
reactions. Low cure stresses are present in the final polymer as there is less volumetric shrinkage and
delayed gelation. The stress-strain responses and tune degradation rates are characterized. Ultimately,
it is envisioned that these unique block copolymer hydrogels will greatly aid in the adhesion,
differentiation and proliferation of cells with potential applications as tissue replacements while
maintaining the targeted mechanical properties.
Title: Multi-Dimensional Biopolymer Embedding Using Custom Built 3D Bio-Printer
By: Tejesh Marsale, Prabir Patra, Usman Halim, Shrishti Singh, Isaac George Macwan, Amer Khamaiseh,
Steven Falzerano
Affiliation: University of Bridgeport
Poster Number: 44
Abstract: The goal of this paper is to demonstrate multidimensional dual extrusion biological printing
with the assistance of a polymer based hydrogel. Current methods of 3D bioprinting are hindered by the
need to create scaffolds to support the intended acellular scaffold which supports embedded cells and
cellular growth. Modern techniques using multiple extruders and angling are limited in their direction of
application and thus their ability to maintain a precise scaffold structure over time. We aim to address
these challenges by constructing and maintaining 3D engineered tissue by a unique robotic 3D printing
method. The printer features a 150μm extruding needle and finely pitched lead screws each run by
finely tuned stepper motors for precision construction. The main printer can be paused, allowing the
rotating plate to be properly angled for the robotic arm to continue embedding, as well as to make
repairs and provide ways to maintain the scaffold from any angle. This new technique uses multiple
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computer aided sliced stereolithographic files to guide the device in overcoming the necessity of
printing a single layer by layer structure at any given angle, allowing for greater control of construction,
maintenance and repair. Using a Cartesian 3D printer in conjunction with a rotating printing platform
and a four axis robotic arm, we have managed to develop a bioprinter capable of overcoming several
limitations facing standard 3D printing techniques. Also, we have produced greater structural support
through the use of a viscous biocompatible polymer substrate and two biological inks, capable of
generating blood vessel scaffold and bone fabrication. The substrate used for the suspension of the bioink consists of hydrogel made from carbopol and gelatin meant for shear elastic modulus and buoyancy
to allow unaffected layer by layer printing.
Title: Patterned polydimethylsiloxane to create micro scaled selective high-surface-energy cell adhesion
zones on polystyrene surface by radio frequency glow discharge treatment
By: Rishab Gaba, Robert Baier
Affiliation: State University of New York at Buffalo
Poster Number: 45
Abstract: A novel technique for developing user-selective cell growth-friendly-regions was developed.
This technique will be of great value for experiments on 3-D printed scaffolds for tissue engineering as it
is independent of biomaterial. A critical observation was made and implored while working on a project
of production of minimally disturbed synchronous fibroblast 3T3 cells. Amidst re-design of an apparatus
called the “baby machine”, it was re-established that just by covering the surface of a bacteria grade,
low-surface-energy polystyrene Petri dish; it could be masked/shielded from the effect of plasma when
subjected to Radio Frequency Glow Discharge Treatment (RFGDT). A layer of polydimethylsiloxane
(PDMS) containing cylindrical holes of 22 µm in diameter and 50 µm in height was fabricated by soft
lithography and replica molding processes. This mask was then hand pressed against the surface of the
Petri dish. This led to creation of an air pocket of cylindrical volume. Experiments involving shear stress
challenges of 3-30 dynes/cm2 to attached cells were conducted to confirm that this air pocket was
sufficient to sustain the glow and hence, selectively treat exposed surfaces. Therefore, differential high
surface energy patterns were generated in congruence with the micro patterns of the PDMS mask. The
high-surface-energy zones, ‘hotspots’ can be of any geometry, size and pattern as the techniques of
generating the patterns on PDMS by soft lithography are fairly advanced when working on a micro scale.
This is possible due to the unique property of radio wave electromagnetic radiations to travel through
any medium. This technique can be exploited to increase cell retention on 3-D printed scaffolds
containing cell ‘compartments’ on biomaterials such as alginate which at present face the obstacle of
poor cell retention.
Title: Passive control of the droplet volume and generation efficiency through the sinusoidal shaped
micro-fluidic channel geometry
By: Bing Han (1), Ruo-Ling Wang (2)
Affiliation: 1) 2011 Co-innovation Center, Nanjing University of Science and Technology, 2) Nanjing
University of Science and Technology
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Poster Number: 46
Abstract: In order to obtain the stable droplet generation with adjustable droplet size and generation
rate, a new kind of channel geometry, i.e. sinusoidal shaped channel, is designed and investigated
numerically in this work. A series of droplets with 30μm-90μm in length, 50μm in width and separated
by 40μm-120μm can be generated at the generation rate from 200/s to 550/s. The generated droplet
size and distance increase and the generation rate decreases, when the geometry of the channel
sinusoidal part becomes more flattened. All kinds of channel geometries are able to generate the
droplets stably for the slip channel-liquid interface, while it is necessary to keep the value of A/λ<=0.14,
where A is the sinusoidal amplitude and λ is the length of the sinusoidal part, to enable the steady
generation of droplets for no-slip channel-liquid interfaces. However, as A/λ decreases beyond 0.14, the
differences in the size of the droplets increase obviously between the two kinds of channel-liquid
interface slippery conditions. Furthermore, a smaller interfacial tension between the dispersed and
continuous phases would enable a higher generation rate of the droplets. This work can be references
to droplet based devices that required for the adjustable dose of the reagent and controllable reaction
rate.
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