Session G21: Novel Instrumentation & Measurements for Biomedical Research

An ultra-low field electron paramagnetic resonance technique for biomedical research

Conventional electron paramagnetic resonance (EPR) imaging systems use high magnetic fields confined to small spaces and therefore have limitations on the sample size and safety issues related to the high level of radiation. We are developing an ultra-low field SQUID (Superconducting Quantum Interference Device)-magnetometer system for EPR detection from room temperature samples in magnetic fields of a few Gauss, corresponding to EPR frequencies of a few MHz. Operation at low EPR excitation frequencies of a few MHz ensures negligible sample heating and high penetration depth in biological systems. A new measurement system consists of a specially designed low noise, non-magnetic 4K dewar with a hollow tail housing a superconducting second order gradiometer inductively coupled to a two-stage dc SQUID amplifier. This unique gradiometer design allows a sample at room temperature to be positioned at the middle turns, which significantly improves the signal-to-noise ratio. We present preliminary results and discuss the prospects for in vivo biomedical EPR imaging.   

A Nanocoaxial-Based Electrochemical Sensor for the Detection of Cholera Toxin

Sensitive, real-time detection of biomarkers is of critical importance for rapid and accurate diagnosis of disease for point of care (POC) technologies. Current methods do not allow for POC applications due to several limitations, including sophisticated instrumentation, high reagent consumption, limited multiplexing capability, and cost. Here, we report a nanocoaxial-based electrochemical sensor for the detection of bacterial toxins using an electrochemical enzyme-linked immunosorbent assay (ELISA) and differential pulse voltammetry (DPV). Proof-of-concept was demonstrated for the detection of cholera toxin (CT). The linear dynamic range of detection was 10 ng/ml - 1 $\mu$g/ml, and the limit of detection (LOD) was found to be 2 ng/ml. This level of sensitivity is comparable to the standard optical ELISA used widely in clinical applications. In addition to matching the detection profile of the standard ELISA, the nanocoaxial array provides a simple electrochemical readout and a miniaturized platform with multiplexing capabilities for the simultaneous detection of multiple biomarkers, giving the nanocoax a desirable advantage over the standard method towards POC applications.   

Symmetric and Asymmetric Split Ring Resonators for Biosensing at Terahertz Frequencies

Food allergies have become a major health concern around the world. Peanut allergies are particularly important because they affect over 5 million people in the United States. We are proposing to develop a metamaterial-based sensor for peanut allergens. The detection mechanism we will tap into is the change in a metamaterial's resonant response due to the presence of a biomolecule in the gap region. Using a commercial-grade simulator based on the finite-difference time-domain method, we have simulated the terahertz transmission and reflection spectra of three different split-ring resonator designs with and without a biomolecule present. By modifying the overall symmetry of the resonator and the geometry of the gap region, we have modified the resonant response and increased its sensitivity. The increased sensitivity is demonstrated by repeating the simulations with a layer of peroxidase conjugated immunoglobulin G (PX-IgG) in the gap region and quantifying the resulting resonant shift. These results are the basis for the proposed allergen sensors.   

On-chip cell sorting via patterned magnetic traps

Due to their importance in research for the diagnosis and treatment of cancer, numerous schemes have been developed to sort rare cell populations, e.g., circulating tumor cells (CTCs), from a larger ensemble of cells. Here, we improve upon a previously developed microfluidic device (Lab Chip 13, 1172, (2013)) to increase throughput and sorting purity of magnetically labeled cells. The separation mechanism involves controlling magnetic forces by manipulating the magnetic domain structures of embedded permalloy microdisks with weak external fields. These forces move labeled cells from the input flow stream into an adjacent buffer flow stream. Such magnetically activated transfer separates the magnetic entities from their non-magnetic counterparts as the two flow streams split apart and move toward their respective outputs. Purity of the magnetic output is modulated by the withdrawal rate of the non-magnetic output relative to the inputs. A proof of concept shows that CTCs from metastatic breast cancer patients can be sorted, recovered from the device, and confirmed as CTCs using separate immunofluorescence staining and analysis. With further optimizations, the channel could become a useful device for high purity final sorting of enriched patient cell samples.   

Neuroelectronic device based on nanocoax arrays

We report on development of a nanocoax-based neuroelectronic array. The device has been used in real time to noninvasively couple to a ganglion sac located along the main nerve cord of the leech $\it{Hirudo}$ $\it{medicinalis}$. This allowed for extracellular recording of synaptic activity in the form of spontaneous synapse firing in pre- and post-synaptic somata, with the next target being recording of local field potentials from rat hippocampal cells. We also discuss an alteration of the architecture to facilitate optical integration of the nanoarray, toward utilizing the so-modified device to elicit / inhibit action potentials in optogenetically-modified cells.   

Intracavity Microfluidic Laser Device for Single Cell Analysis

An intracavity microfluidic laser device has been developed to study bioparticles ranging in size from 50 nm to 20 $\mu$m (virons to organelles to whole cells). The versatile device can be operated used in several modes including static or flowing fluids, with or without molecular labels, and microscopic imaging and/or spectroscopy. It enables advantageous new ways to perform analyses of bioparticles for applications including cell biology, detection of disease and pathogens, environmental monitoring, pharmaceuticals, agriculture, and food processing. This talk will briefly summarize the physics of the device including its laser optics, fluid dynamics, and intracavity light interaction with cells. The talk will then focus on results of a study of mitochondria in normal and cancer liver cells. The study examines the transformation of intracellular and isolated mitochondria from the normal to disease state. The results highlight the unique utility of the device to rapidly assess biophysical changes arising from altered biomolecular states of cells and organelles.   

Individual Mammalian Cell Magnetic Measurements with a Superconducting Quantum Interference Device

Magnetism can be introduced into otherwise nonmagnetic cells by the uptake of superparamagnetic iron oxide (SPIO) nanoparticles. SPIO nanoparticles are used in numerous biomedical applications including cellular therapies and targeted drug delivery. Currently there are few tools capable of characterizing individual magnetic nanoparticles and the magnetic properties of individual mammalian cells loaded with SPIO. Our scanning superconducting quantum interference devices (SQUIDs) are good candidates for these measurements due to their high sensitivity to magnetic dipole moments (approx. 200 $\mu_{\mathrm{b}}$/$\surd $Hz) In this study, we use a scanning SQUID to image the magnetic flux from SPIO loaded H1299 lung cancer cells. We find that the magnetic moment spatially varies inside the cell with each cell having a unique distribution of moments. We also correlate these magnetic images with optical and scanning electron microscope images. These results show that the SQUID is a useful tool for imaging biological magnetism. The visualization of single cell magnetism and the quantification of magnetic dipole moments in magnetically labeled cells can be used to optimize conventional biological magnetic imaging techniques, such as MRI.   

Interaction-Free Quantum Electron Microscope in Free-Space

We propose the design and theoretical analysis of a quantum electron microscope (QEM), which utilizes interaction-free quantum measurement with electrons for nanoscale imaging. The QEM can be used to image electron-irradiation-sensitive materials, such as biological samples, with a high resolution and low radiation damage. Our QEM scheme is an electron interferometer with a storage resonator. The incoming electron beam is asymmetrically split into a strong reference beam and a weak sample beam, both of which are stored in the resonator. Only the weak sample beam transmits through the sample for multiple times. We propose to build the QEM with free-space electron optics. We develop a scattering matrix method to theoretically analyze the contrast mechanism, radiation damage, and measurement accuracy. We propose an electron-mirror-based storage resonator and we have performed electron optics simulation of electron trajectories within the resonator. We also report experimental implementation and characterization of the electron beam-splitter to be used in the QEM. Thin crystals fabricated with focused ion beam and nano-gratings fabricated with electron-beam lithography are two candidate beam-splitters, both of which are characterized by electron diffraction.   

Light localization properties of biological cells via confocal imaging

Detection and characterization of the spatial refractive index fluctuations of very weakly disordered optical dielectric media has ample applications in various fields ranging from soft condensed matter to biological research. We report a study of the submicron scale degree of the structural disorder of heterogeneous weakly disordered optical dielectric media, such as biological cells, by quantifying their submicron scale light-localization properties. Confocal microscopy is used to construct disordered optical lattices of these dielectric media. Light-localization properties are studied by the statistical analysis of the inverse participation ratio (IPR) of the localized eigenfunctions of these optical lattices at the submicron scales. The method is described and its importance is highlighted. As one of the applications, we demonstrate that using this method, different types of normal and cancerous cells can be distinguished by quantifying the structural disorder inside the cells via their confocal micrographs. Other potential applications of the technique to characterize weakly disordered media, as well as biological cells, in particular cancer detection, are also discussed.   

Enhanced partial wave spectroscopy (EPWS) for nanoscale sensitive structural disorder measurement in weakly disordered media

Based on mesoscopic physics approach, partial wave spectroscopy (PWS) technique was introduced earlier for the measurement of the nanoscale structural disorder in very weakly disordered optical media, such as biological cells. We describe a modification and further improvement of this technique, by introducing an enhanced back reflection system to further increase the measurement sensitivity of the PWS technique. As a result, a nanoscale level of fluctuation or nanoscale alteration in a disordered medium can be measured with even higher sensitivity and accuracy. In this enhanced-PWS (EPWS) technique, semi-transparent metallic thin films are used to create a cavity like structure that holds the sample; this leads to an increased back reflected light intensity, which eventually results in high sensitivity in the nanoscale structural disorder measurements. Reflection coefficients of the backscattered signal obtained from the simulated disordered media, within the cavity, with varying refractive index fluctuation values, were statistically analyzed using mesoscopic physics approach. The results, of a sample with and without metallic cavity inside the system show a significantly high back reflected intensity with the metallic cavity case. We also discuss the possible applications of the developed technique in ultra-sensitive detection of cancer by the characterization of nanoscale fluctuations in biological cells.   

Quantitative ultrasonic bone assessment using backscatter measurements at 1 MHz

Osteoporosis is a complicated metabolic degenerative bone disease affecting millions of Americans. The current standard for diagnosis is an x-ray technique called DXA, but ultrasound may offer a cheaper, more portable diagnosis method that may also be sensitive to bone strength in ways DXA is not. Ultrasonic backscatter techniques in the 5 MHz range have been shown to be sensitive to changes in bone mineral density (BMD) associated with osteoporosis. These techniques measure the power ratio between an earlier and later part of the backscatter signal. These ratios (called nMBD in the frequency domain or nBAR in the time domain) depend on ultrasonic attenuation in bone which is known to correlate with BMD. This study measured nMBD and nBAR using a 1 MHz transducer. Lower frequency transducers increase signal penetration into bone. Linear regression analysis was used to investigate correlations between nMBD and nBAR and the density and microstructural characteristics of bone such as the structural model index (SMI) and trabecular number (TbN). Good linear correlations were observed for nMBD and nBAR vs. BMD at r$=$0.75 and r$=$0.70, respectively, comparable to correlations obtained using a 5 MHz transducer. Correlations with SMI and TbN were within the r$=$0.65-0.75 range.   

Thermal (rf) and non-thermal mechanisms of nanoparticles induced/enhanced cancer cell apoptosis

It was demonstrated that the \textit{rf} procedures can be non-invasive and cancer selective when combined with nanoparticles (NPs) that work as \textit{rf} heating enhancers. However, there are disparities, between theory and experimental results, especially for non-magnetic NP. Therefore, it is necessary to elucidate the physical mechanisms that control the reported \textit{rf} heating. We have constructed an apparatus for \textit{rf} heating, which allows for applying either E$_{\mathrm{rf}}$ or H$_{rf}$ fields in the kHz-MHz frequency range. Our results of specific absorption rate (SAR) measurements for both magnetic and nonmagnetic of NPs indicate that \textit{rf} electric field also plays the role in heating of magnetic NPs and that in the nonmagnetic case only interface losses are responsible for the observed heating. In search for a more efficient and non-thermal method, we have explored a cancer cell death through mechanical stress imposed on the cell membrane. We have designed a special setup to apply either static or ac magnetic fields/gradients (up 300T/m) to cultured cancer cell lines with/without PNs added. The fields and gradients, and forces applied were simulated using HFSS/Maxwell software. Pancreatic adenocarcinoma cell line, AsPC-1 stained with DRAQ7 were studied. Very strong dependence of number of dead cells on applied field strength was observed. Discussion of the two mechanisms (\textit{rf} and non-\textit{rf}) of observed apoptosis will be presented.   

Precise control of DNA recapture in solid-state nanopore

Solid-state nanopore is a novel experimental method in detecting and analyzing single biomolecules such as DNA, protein and virus. The dynamic behavior of such molecules in a microfluidic system can be investigated by the back-and-forth translocation control. Such motion control is made possible by fast changing of the polarity of the driving voltage, and the repeat measurement of a single molecule is expected to increase the signal-to-noise-ratio (SNR) significantly. However, due to the existence of membrane capacitance and electrolyte resistance, transient current spike raises as driving voltage changes. Such current spike saturates the data acquisition system and leads to difficulty in detecting fast returned molecules. Simulation shows that the electric field in electrolyte is proportional to the ionic current and increases dramatically during the transient charging period. This extra electric force pulls the molecule back to the pore faster than expected. Our study demonstrates that the transient current can be compensated by modifying the profile of the driving voltage. With such improvement, the observed distribution of recaptured translocation events matches perfectly with the prediction by drift-diffusion transport equation. This finding will help building precise control technique towards DNA sequencing in nanopore.   

Assessing the performance of trans rectal ultrasound to measure prostate weight and its association with other diagnostic factors

Trans rectal ultrasound is fast becoming an important tool used in the prognosis of prostate cancer. But how does it compare to other measures, such as the actual weight of the prostate obtained after radical prostatectomy. Here, we assess the association of prostate weight obtained via trans rectal ultrasound and actual specimen weight obtained after radical prostatectomy with body mass index (BMI) using linear regression models adjusted for Gleason score, pre-operative PSA, and age applied to subsets of Euro-Americans (n $=$ 242) and African Americans (n $=$ 34). The roles of both prostate weight and BMI are themselves part of ongoing research focused on prostate cancer prognosis. Our preliminary results show a marginal relationship between BMI and specimen weight obtained after surgery in Euro-Americans but no relationship between BMI and ultrasound measured weight in either race subset. Therefore, further work pertaining to the performance of trans rectal ultrasound may be warranted. This work is supported by the Northwestern University SPORE in Prostate Cancer (NIH/NCI P50 CA 90386).   

How to get into that ``room at the bottom'' of DNA analysis

Linearly extending long DNA molecules in sub-50 nm nanochannels for genomic analysis, while retaining their structural integrity, is a major technological challenge. We employ ``Convex Lens-induced Confinement'' (CLiC) microscopy to gently load DNA into nanogrooves from above, overcoming the limitations of side-loading techniques used in direct-bonded nanofluidic devices. In the CLiC technique, the curved surface of a convex lens is used to deform a flexible coverslip above a glass substrate, creating a nanoscale gap that can be tuned during an experiment to load and confine molecules into nanoscale features embedded in the bottom substrate. Since DNA molecules are loaded into the embedded nanotopography from above, CLiC eliminates the need for the high pressures or electric fields required to load DNA into direct-bonded nanofluidic devices. To demonstrate the versatility of CLiC, we confine DNA to a variety of nanostructures, demonstrating DNA nanochannel-based stretching and denaturation mapping. In particular, we have successfully extended DNA in 27 nm channels, achieving high stretching (90 percent) that is in good agreement with Odijk deflection theory, and we have mapped genomic features using denaturation analysis.