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This PDF file contains the front matter associated with SPIE Proceedings Volume 8585, including the Title Page, Copyright information, Table of Contents, and the Conference Committee listing.
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Microfluidic devices have been widely used in manipulation and analysis of individual cells in small-volume solutions. It
could be potentially used for studies of the interaction of THz radiation with biomolecules and cells in aqueous media.
We present a prototype microfluidic device that can be used for controlled cellular exposures to THz radiation. The
device is made of a PDMS microfluidic channel on glass substrate and consists of electrodes for cell concentration.
Initial cell concentration and THz transmission measurements have been performed on various prototype samples. Our
results demonstrate the feasibility of using microfluidic chips for potential “Lab-on-a-Chip” THz applications.
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Many applications of terahertz technology are concerned with sensing of substances such as drugs, chemical compounds,
explosives and much more. For this purpose, low-cost terahertz measurement systems are desired. In this respect, metasurfaces
can be used as sensitive near-field sensors by exploiting the change of resonant frequency in the vicinity of
substances. We demonstrate chip-based terahertz sensors that can be applied to measure the thickness of ultra-thin
materials with a resolution of the order of 1/16000 of the wavelength. Furthermore, we show that the same sensor can be
used for refractometric measurements. In this context, we evaluated the refractive index of highly absorptive liquids and
liquid mixtures. Based on these measurements, we retrieved the mixing ratio of the liquid mixtures. Moreover, we show
that meta-surfaces can be employed to implement chip-based terahertz circuits for highly confined surface waves. The
electromagnetic properties of the meta-surface can be designed on purpose. For example, such meta-surfaces can serve
as integrated interferometric sensors and can be used for highly sensitive measurements when only a small amount of a
sample material is available.
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The interfaces of a dielectric sample are resolved in reflection geometry using light from a frequency agile array of
terahertz quantum-cascade lasers. The terahertz source is a 10-element linear array of third-order distributed feedback
QCLs emitting at discrete frequencies from 2.08 to 2.4 THz. Emission from the array is collimated and sent through a
Michelson interferometer, with the sample placed in one of the arms. Interference signals collected at each frequency are
used to reconstruct an interferogram and detect the interfaces in the sample. Due to the long coherence length of the
source, the interferometer arms need not be adjusted to the zero-path delay. A depth resolution of 360 μm in the
dielectric is achieved with further potential improvement through improved frequency coverage of the array. The entire
experiment footprint is <1 m x 1 m with the source operated in a compact, closed-cycle cryocooler.
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We report a method of taking mid-infrared and terahertz spectra on nanoscale using compact mW-level sources, such as quantum cascade lasers, and a standard atomic force microscope (AFM). Light absorption is detected via deflection of an AFM cantilever due to local sample thermal expansion. The spatial resolution is principally determined by the diameter of the high-intensity spot in the vicinity of a sharp metalized AFM tip, and is below 50nm. To enable detection of minute sample expansion, the repetition rate of the laser pulses is moved in resonance with the cantilever mechanical frequency. The technique requires no optical detectors.
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Structural modifications of cell membranes are among the primary consequences of exposure to intense nanosecond
pulsed electric fields. These alterations can be characterized indirectly by monitoring changes in electrical conductance
or small molecule permeability of artificial membranes or suspensions of living cells, but direct observations of the
membrane-permeabilizing structures remain out of the reach of experiments. Molecular dynamics simulations provide an
atomically detailed view on the nanosecond time scale of the sequence of events that follows the application of an
external electric field to a system containing an aqueous electrolyte and a phospholipid bilayer, a simple approximation
of a cell membrane. This biomolecular perspective, which correlates with experimental observations of electroporation
(electropermeabilization) in many respects, points to the key role of water dipoles, driven by the electric field gradients
at the membrane interface, in the initiation and construction of the membrane defects which evolve into conductive
pores. We describe a method for stabilizing these lipid electropores in phospholipid bilayers, and for characterizing their
stability and ion conductance, and we show how the properties of these nanoscale structures connect with continuum
models of electroporation and with experimental results.
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For long, the contribution of water network motions to enzymatic reactions was enigmatic due to the complexity of biological systems and to experimental limitations. Thanks to the development of new powerful THz emitters and detectors in the last decades, it is now possible to probe dynamics on the timescale of the fast hydrogen bond rearrangements during biochemical reactions. For this purpose, we developed a kinetic terahertz absorption (KITA) spectrometer which combines the strength of THz radiation (~1012 Hz = 1 ps) to directly probe collective picosecond protein-water dynamics with the fast mixing properties of a stopped-flow apparatus which initializes a biochemical reaction within milliseconds. With KITA, we analyzed the collective water dynamics during substrate hydrolyses by a human matrix-metalloproteinase. In addition, we studied the reorganization and electrostatic changes at the catalytic zinc-ion from the enzyme active site and performed molecular dynamics simulations of the enzyme-substrate-water system. Our results revealed a systematic gradient of water network motions: From the active site to the bulk water hydrogen bond dynamics increased from 7 ps (active site) to 1ps (bulk water) prior to substrate binding and hydrolysis. The approaching substrate perturbs the dynamic water gradient resulting in an overshoot of KITA signal which then relaxes back during onset of substrate hydrolyses. Our findings suggest that collective water dynamics may contribute to effective substrate binding to enzyme active sites and could be induced by the charge of the catalytic zinc-ion residing at the active site.
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Terahertz time-domain spectroscopy (THz-TDS) methods have been utilized in previous studies in order to characterize
the optical properties of skin and its primary constituents (i.e., water, collagen, and keratin). However, similar
experiments have not yet been performed to investigate whether melanocytes and the melanin pigment that they
synthesize contribute to skin’s optical properties. In this study, we used THz-TDS methods operating in transmission
geometry to measure the optical properties of in vitro human skin equivalents with or without normal human
melanocytes. Skin equivalents were cultured for three weeks to promote gradual melanogenesis, and THz time domain
data were collected at various time intervals. Frequency-domain analysis techniques were performed to determine the
index of refraction (n) and absorption coefficient (μa) for each skin sample over the frequency range of 0.1-2.0 THz. We
found that for all samples as frequency increased, n decreased exponentially and the μa increased linearly. Additionally,
we observed that skin samples with higher levels of melanin exhibited greater n and μa values than the non-pigmented
samples. Our results indicate that melanocytes and the degree of melanin pigmentation contribute in an appreciable
manner to the skin’s optical properties. Future studies will be performed to examine whether these contributions are
observed in human skin in vivo.
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The use of nanosecond pulsed electric fields to ablate tumors (nanoelectroablation) is now well established in the murine xenograft model system. In order bring this therapy into the clinic for the treatment of human tumors we are developing both a pulse generator as well as delivery electrodes to target the tumors to be treated. We will describe the PulseCure® Model MBR-1 100 ns pulse generator and the first human clinical trial data using nanoelectroablation to scarlessly ablate basal cell carcinomas.
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Non-compressible hemorrhages are the most common preventable cause of death on battlefield or in civilian traumatic
injuries. We report the use of sub-millisecond pulses of electric current to induce rapid constriction in femoral and
mesenteric arteries and veins in rats. Extent of vascular constriction could be modulated by pulse duration, amplitude
and repetition rate. Electrically-induced vasoconstriction could be maintained at steady level until the end of
stimulation, and blood vessels dilated back to their original size within a few minutes after the end of stimulation. At
higher settings, a blood clotting could be introduced, leading to complete and permanent occlusion of the vessels. The
latter regime dramatically decreased the bleeding rate in the injured femoral and mesenteric arteries, with a complete
hemorrhage arrest achieved within seconds. The average blood loss from the treated femoral artery was about 7 times
less than that of a non-treated control. This new treatment modality offers a promising approach to non-damaging
control of bleeding during surgery, and to efficient hemorrhage arrest in trauma patients.
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Terahertz time-domain spectroscopy (THz-TDS) systems are capable of detecting small differences in water
concentration levels in biological tissues. This feature makes THz devices excellent tools for the noninvasive assessment
of skin; however, most conventional systems prove too cumbersome for limited-space environments. We previously
demonstrated that a portable, compact THz spectrometer permitted measurement of porcine skin optical properties that
were comparable to those collected with conventional systems. In order to move toward human use of this system, the
goal for this study was to collect the optical properties, specifically the absorption coefficient (μa) and index of refraction
(n), of human subjects in vivo. Spectra were collected from 0.1-2 THz, and measurements were made on the palm,
ventral (inner) and dorsal (outer) forearm. Prior to each THz measurement, we used a multiprobe adapter system to
measure each subject’s skin hydration levels, transepidermal waterloss (TEWL), skin color, and degree of melanin
pigmentation. Our results suggest that the measured optical properties were wide-ranging, and varied considerably for
skin tissues with different hydration and melanin levels. These data provide a novel framework for accurate human
tissue measurements using THz spectrometers in limited-space environments.
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In recent years, many applications have been recognized for biomedical imaging techniques utilizing terahertz frequency
radiation. This is largely due to the capability of unique tissue identification resulting from the nature of the interaction
between THz radiation and the molecular structure of the cells. By THz identification methods, tissue changes in tooth
enamel, cartilage, and malignant cancer cells have already been demonstrated. Terahertz Time-Domain Spectroscopy
(THz-TDS) remains one of the most versatile methods for spectroscopic image acquisition for its ability to
simultaneously determine amplitude and phase over a broad spectral range.
In this study we investigate the use of THz imaging techniques to uniquely identify damage types in tissue samples for
both forensic and treatment applications. Using THz-TDS imaging in both transmission and reflection schemes, we
examine tissue samples which have been damaged using a variety of acids. Each method of damage causes structural
deterioration to the tissue by a different mechanism, thus leaving the remaining tissue uniquely changed based on the
damage type. We correlate the change in frequency spectra, phase shift for each damage type to the mechanisms and
severity of injury.
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We have for the first time recorded action potentials in rat hippocampus neurons when they were stimulated by
subnanosecond electric pulses. The preliminary results show that applying a series of pulses allowed the accumulation of
depolarization before activating the voltage gated channels. The depolarization only occurred when the electric pulses
were applied. It is unclear whether the depolarization is caused by the charge accumulation across the membrane or the
cation influx due to the membrane permeabilization. We have also conducted an electromagnetic simulation of
delivering subnanosecond pulses to tissues using an impulse radiating antenna. The results show that the pulses can be
confined in the deep region in the brain but the amplitude is reduced significantly due to the attenuation of the tissues. A
partially lossy dielectric lens may be used to reverse the decreasing trend of the electric field.
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The use of electrically-induced neuromodulation has grown in importance in the treatment of multiple neurological
disorders such as Parkinson’s disease, dystonia, epilepsy, chronic pain, cluster headaches and others. While electrical
current can be applied locally, it requires placing stimulation electrodes in direct contact with the neural tissue. Our goal
is to develop a method for localized application of electromagnetic energy to the brain without direct tissue contact.
Toward this goal, we are experimenting with the wireless transmission of millimeter wave (MMW) energy in the 10-100
GHz frequency range, where penetration and focusing can be traded off to provide non-contact irradiation of the cerebral
cortex. Initial experiments have been conducted on freshly-isolated leech ganglia to evaluate the real-time changes in the
activity of individual neurons upon exposure to the MMW radiation. The initial results indicate that low-intensity
MMWs can partially suppress the neuronal activity. This is in contrast to general bath heating, which had an excitatory
effect on the neuronal activity. Further studies are underway to determine the changes in the state of the membrane
channels that might be responsible for the observed neuromodulatory effects.
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Exposure to nano-second pulsed electrical fields (nsPEFs) has been shown to cause poration of external and internal cell
membranes, DNA damage, and blebbing of the plasma membrane. Recovery from nsPEF exposure is likely dependent on
multiple factors, including exposure parameters, length of time between pulses, and extent of cellular damage. As cells
progress through the cell cycle, variations in DNA and nucleus structure, cytoskeletal arrangement, and elasticity of cell
membrane could cause nsPEFs to affect cells differently during different cell cycle phases. To better understand the
impact of nsPEF on cell cycle, we investigated CHO cell cycle progression following varying intensities of nsPEFexposures.
Cell populations were examined post exposure (10 ns pulse trains at 100, 150, or 200kV/cm) by analysis of
DNA content via propidium iodide staining and flow cytometric analysis to determine cell cycle phase. Populations
exhibited arrest in G2/M phase, but not in G1 phase at 1h post-exposure that increased in severity and duration with
increasing exposure dose. Recovery from arrest was complete after 12h, and populations did not exhibit an increase in
apoptosis as a result of exposure. Post exposure arrest in G2/M phase may indicate that nsPEF-induced damage is not
significant to cause G1 arrest or that mitotic checkpoints are more important regulators of cell cycle progression after
nsPEF exposure.
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Pulsed terahertz (THz) imaging has been suggested as a novel high resolution, noninvasive medical diagnostic tool.
However, little is known about the influence of pulsed THz radiation on human tissue, i.e., its genotoxicity and effects on
cell activity and cell integrity. We have carried out a comprehensive investigation of the biological effects of THz
radiation on human skin tissue using a high power THz pulse source and an in vivo full-thickness human skin tissue
model. We have observed that exposure to intense THz pulses causes DNA damage and changes in the global gene
expression profile in the exposed skin tissue. Several of the affected genes are known to play major roles in human
cancer. While the changes in the expression levels of some of them suggest possible oncogenic effects of pulsed THz
radiation, changes in the expression of the other cancer-related genes might have a protective influence. This study may
serve as a roadmap for future investigations aimed at elucidating the exact roles that all the affected genes play in skin
carcinogenesis and in response to pulsed THz radiation.
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Application of nanosecond pulsed electric fields (nsPEF) to various biological cell lines has been to shown to cause
many diverse effects, including poration of the plasma membrane, depolarization of the mitochondrial membrane,
blebbing, apoptosis, and intracellular calcium bursts. The underlying mechanism(s) responsible for these diverse
responses are poorly understood. Of specific interest in this paper are the long-term effects of nsPEF on cellular
processes, including the regulation of genes and production of proteins. Previous studies have reported transient
activation of select signaling pathways involving mitogen-activated protein kinases (MAPKs), protein
phosphorylation and downstream gene expression following nsPEF application. We hypothesize that nsPEF
represents a unique stimulus that could be used to externally modulate cellular processes. To validate our
hypothesis, we performed a series of cuvette-based exposures at 10 and 600ns pulse widths using a custom Blumlien
line pulser system. We measured acute changes in the plasma membrane structure using flow cytometry by tracking
phosphatidylserine externalization via FITC-Annexin V labeling and poration via propidium iodide uptake. We then
compared these results to viability of the cells at 24 hours post exposure using MTT assay and changes in the
MAPK family of proteins at 8 hours post-exposure using Luminex assay. By comparing exposures at 10 and 600ns
duration, we found that most MAPK family-protein expression increased in Jurkat and U937 cell lines following
exposure and compared well with drops in viability and changes in plasma membrane asymmetry. What proved
interesting is that some MAPK family proteins (e.g. p53, STAT1), were expressed in one cell line, but not the other.
This difference may point to an underlying mechanism for observed difference in cellular sensitivity to nsPEFinduced
stresses.
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The exposure of nanosecond pulsed electric fields (nsPEF) to living cells has been shown to create nanopores in the
plasma membranes. These nanopores allow the passage of small ions but exclude the transport of larger molecules
such as Propidium ions, with permeabilization persisting for many minutes. To characterize these nanopores and the
effect of temperature of the formation and resealing of these pores, we have chosen to use 6-Propionyl-2-(N,NDimethylamo)
Naphthalene (PRODAN) as an indicator of membrane organization. PRODAN is a fluorescent dye
with a large excited-state dipole moment that displays extensive solvent polarity-dependent fluorescent shifts. By
monitoring this shift in fluorescence spectrum, disruption of the membrane after an electric exposure is observed as
an immediate increase in the membrane fluidity, likely indicating poration of the membrane. High-speed imaging
results indicate that a change in membrane organization occurs instantly (<5 ms), with longer pulse widths having a
more dramatic effect on the membrane. This instantaneous membrane disruption was shown to recover within 500
ms.
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Nanosecond pulsed electric fields (nsPEFs) are known to increase cell membrane permeability to small molecules in
accordance with dosages. As previous work has focused on nsPEF exposures in whole cells, electrodeformation may
contribute to this induced-permeabilization in addition to other biological mechanisms. Here, we hypothesize that
cellular elasticity, based upon the cytoskeleton, affects nsPEF-induced decrease in cellular viability. Young’s moduli of
various types of cells have been calculated from atomic force microscopy (AFM) force curve data, showing that CHO
cells are stiffer than non-adherent U937 and Jurkat cells, which are more susceptible to nsPEF exposure. To distinguish
any cytoskeletal foundation for these observations, various cytoskeletal reagents were applied. Inhibiting actin
polymerization significantly decreased membrane integrity, as determined by relative propidium uptake and
phosphatidylserine externalization, upon exposure at 150 kV/cm with 100 pulses of 10 ns pulse width. Exposure in the
presence of other drugs resulted in insignificant changes in membrane integrity and 24-hour viability. However, Jurkat
cells showed greater lethality than latrunculin-treated CHO cells of comparable elasticity. From these results, it is
postulated that cellular elasticity rooted in actin-membrane interaction is only a minor contributor to the differing
responses of adherent and non-adherent cells to nsPEF insults.
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Terahertz (THz) hydration sensing continues to gain traction in the medical imaging community due to its unparalleled
sensitivity to tissue water content. Rapid and accurate detection of fluid shifts following induction of thermal skin burns
as well as remote corneal hydration sensing have been previously demonstrated in vivo using reflective, pulsed THz
imaging. The hydration contrast sensing capabilities of this technology were recently confirmed in a parallel 7 Tesla
Magnetic Resonance (MR) imaging study, in which burn areas are associated with increases in local mobile water
content. Successful clinical translation of THz sensing, however, still requires quantitative assessments of system
performance measurements, specifically hydration concentration sensitivity, with tissue substitutes. This research aims
to calibrate the sensitivity of a novel, reflective THz system to tissue water content through the use of hydration
phantoms for quantitative comparisons of THz hydration imagery.Gelatin phantoms were identified as an appropriate
tissue-mimicking model for reflective THz applications, and gel composition, comprising mixtures of water and protein,
was varied between 83% to 95% hydration, a physiologically relevant range. A comparison of four series of gelatin
phantom studies demonstrated a positive linear relationship between THz reflectivity and water concentration, with
statistically significant hydration sensitivities (p < .01) ranging between 0.0209 - 0.038% (reflectivity: %hydration). The
THz-phantom interaction is simulated with a three-layer model using the Transfer Matrix Method with agreement in
hydration trends. Having demonstrated the ability to accurately and noninvasively measure water content in tissue
equivalent targets with high sensitivity, reflective THz imaging is explored as a potential tool for early detection and
intervention of corneal pathologies.
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Terahertz (THz) sensing has shown potential as a novel imaging modality in medical applications due to
its high water sensitivity. The design of medical THz sensing systems and their successful application to
in vivo settings has attracted recent interest to the field, and highlighted the need for improved
understanding of the interaction of THz waves with biological tissues. This paper explores the modeling
of composite materials which combine strongly-interacting water with weakly-interacting species such as
those that are common to biological tissues. The Bruggeman, Maxwell-Garnett, and power law effective
media models are introduced and discussed. A reflection-mode 100 GHz Gunn diode sensing system was
used to measure the reflectivity of solutions of water and dioxane as a function of relative concentration,
and the results were compared with the predictions of the Maxwell-Garnett, power law, and Bruggeman
mixing theories. The Maxwell-Garnett model fit poorly to experimental data on near-equal mixtures of
water and dioxane and improved when the concentration of water exceeded ~55% or was below ~15%.
The first-order power law model fit poorly to experimental data across the entire range except at nearpure
solutions. Power law models employing 1/2 and 1/3 terms improved goodness of fit, but did not
match the accuracy of the Bruggeman model. The Bruggeman provided the best fit to experimental data
model as compared to Maxwell-Garnett and the power models and accurately predicted the solution
reflectivity through the whole range of concentrations. This analysis suggests that the Bruggeman model
may offer improved accuracy over more conventional dielectric mixing models when developing
simulation tools for THz reflectometry of hydrated biological tissues.
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Devices operating at THz frequencies have been continuously expanded in many areas of application and major research
field, which requires materials with suitable electromagnetic responses at THz frequency ranges. Unlike most naturally
occurring materials, novel THz metamaterials have proven to be well suited for use in various devices due to narrow and
tunable operating ranges. In this work, we present the results of two THz metamaterial absorber structures aiming two
important device aspects; polarization sensitivity and broad band absorption. The absorbers were simulated by finite
element method and fabricated through the combination of standard lift-off photolithography and electron beam metal
deposition. The fabricated devices were characterized by reflection mode THz time domain spectroscopy. The narrow
band absorber structures exhibit up to 95% absorption with a bandwidth of 0.1 THz to 0.15 THz.
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Terahertz based spectroscopy and imaging has become an active field of research in the past decade for a plethora of
applications including security screening, biomedical imaging, chemical analysis, and investigation of carrier dynamics.
Several advantages exist for the use of THz techniques since investigation of a sample can be performed without contact
or ionization; however, fine detail is difficult to determine due to the diffraction limit of the radiation. The resolution
limit of THz imaging and sensing can be overcome by the incorporation of near-field optical techniques; which can
allow image resolution as fine as tens of nanometers at THz frequencies. With this expanded resolution capability, THz
imaging can decipher micro- and nano-structural information which, when coupled with the non-contact features of
these techniques, makes THz spectroscopy ideal for the analysis micro and nano-optical devices. In this study, we
demonstrate the development and performance of an aperture-less near-field system which has been integrated to
perform highly-spatially resolved Terahertz Time-Domain Spectroscopic (THz-TDS) imaging.
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