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TRACK 6 TRACK 6 Technical Program

which is more specific. Besides, our nanoparticle-based detection assay is this result with a computational model of a stepwise binding mechanism.
fast and easy to perform and could become the new gold standard for clini- Second, we found the strong binding ligand (fucosyl-GM1) could activate the
cal diagnosis of infectious diarrhea. very weak binding ligand (GM2). A fucosyl-GM1/GM2 mixture increased the
maximum binding of CTB on membrane surfaces. As such, the attenuation
or enhancement of CTB binding is not simply controlled by the concentra-
Micro-Sample Exosome Detection Assay For Pancreatic Cancer tion of strong ligands; the cooperative actions among gangliosides in a com-
plex can influence the overall binding. These unexpected discoveries not
only demonstrate the essence of cooperativity in the multivalent lectin bind-
Poster Presentation. NEMB2016-6086 ing, but also significantly impact lectin-based glycomic analysis. In summary,
our nanocube-based cell membrane array provides an excellent tool to
Dali Sun, Kai Liang, Houston Methodist Research Institute, Hous- dissect complex multivalent interactions; its easy-to-use and high-throughput
ton, TX, United States, Tony Y. Hu, Houston Methodist Research features will make this tool immediately available to biological communities.
Insititute, Houston, TX, United States

Exosome have gained increasing interest as a novel target for cancer di- A fully integrated impedance biosensing platform for point-of-
agnostics. However, detection of tumor-derived exosomes is technically care diagnostics
challenging and often requires extensive sample purification from human
serum. Here we developed an ultrasensitive plasmonic scattering assay that Poster Presentation. NEMB2016-6012
could directly level tumor-derived exosome from down to 1 µl serum. Our
strategy integrated specific three-probe recognition system and plasmonic
coupling effect colorimetric scatter, caused by the proximity of diverse gold Tae-Hoon Kim, Jungkyu Kim, Texas Tech University, Lubbock, TX,
nanoparticles, to enhance local signal intensity of scattering light in dark United States
field microscope. By incorporating tumor-identified maker EphA2, we could
distinguish pancreatic cancer patients from pancreatitis and normal controls An impedance biosensor is a well-known label-free system simplifying
with high specificity. Quantification of circulating tumor-derived exosome overall bioassay procedures. However, the readout sensitivity afforded by
was informative in staging tumor progression and patients’ early response to label-free affinity impedance biosensors is inferior to other label-free tech-
neoadjuvant therapy. niques such as conventional ELISAs. Micro/nanoparticles are a common sig-
nal amplification technique to improve the sensitivity of an impedance based
biosensing platform. However, utilizing the micro/nanoparticle increases
A binding cooperativity study of cholera toxin-mixed ganglio- complexity of overall assay procedures. In this work, we developed a fully
sides using a high-throughput nanocube-based cell membrane integrated impedance biosensing platform which enables high sensitivity
improving SNR (signal to noise ratio) with a microparticle. Also, we incorpo-
array rated a particle labeling method to further improve sensitivity in impedimet-
ric bioassay, and evaluated the minimum detectable number of polymeric
Poster Presentation. NEMB2016-6093 microparticles on an interdigitated electrode (IDE) surface over a wide range
of frequencies using the developed impedance analyzing platform.
Hung-jen Wu, Nolan Worstell, Pratik Krishnan, Joshua Weather-
ston, Texas A&M University, College Station, TX, United States A custom-made impedance measurement system consists of four major
parts: 1) a gold (Au) IDE array chip, 2) an impedance analyzing circuit, 3) a
Lectins often consist of multiple binding subunits that exhibit specific or data acquisition (DAQ) board and 4) a LabVIEW software. The Au IDE array
semi-specific glycan recognition. The cooperative action between multiple chip having a finger width and spacing of 10 um was fabricated on a glass
bound receptors often strongly enhances the binding avidity and specificity. wafer, and 4 sets of IDEs were aligned on the single chip. The surface of
Although the binding subunits of lectins are often identical and many lectins each IDE was functionalized with anti-TNF alpha capture antibody using a
preferentially bind to the same glycan structure, they still exhibit unique standard silane functionalization and bio-conjugation methods. For a sam-
binding patterns to various cell surfaces. We hypothesize that the unique ple handling processor, Lifting Gate microfluidic valves were developed for
lectin binding patterns on heterogeneous cell surfaces are achieved via co- delivering all required samples and reagents, and washing non-specifically
operative interaction between bound glycan moieties. bound targets and microparticles during this immunoassay. As a labeling, a
streptavidin-coated polystyrene (PS) particle was used for impedance mea-
To better understand the essential nature of binding cooperativity in multi- surement. Also, the impedance signal from solely distilled (DI) water was uti-
valent binding mechanism, quantitative analysis of multivalent membrane lized as the baseline. After adding the microparticles with different concen-
recruitment onto the cellular surface is critical. We have developed a trations through the pumping system, the magnitude of impedance signals
nanocube sensor coupled with complex reaction analysis to quantitatively was measured in the specified range of frequency (11 kHz to 91 kHz).
explore the multivalent binding mechanism. The nanocube sensor is sur-
rounded by a lipid bilayer that possesses the same physical and chemical We found a significant decrease of impedance magnitude as the number
properties as cell membranes. This novel sensor is an ideal tool for studying of microparticle increases on the working electrode when compared with
binding cooperativities because receptors can freely diffuse and rotate on label-free approach. Considering the surface coverage rate, the absolute
2D fluidic cellular membranes allowing receptor self-organization to enable magnitude of impedance showed a significant difference with microparticle
multivalent interactions. This label-free sensing platform can be conducted numbers. This result shows a promising possibility of the developed imped-
in standard 384-well microplate; therefore, its high-throughput utility enables ance system in differentiating the amount of surface charge for unknown
the complex analysis of multivalent lectin binding. In addition, the simple microparticles. This fact can be directly related to selectivity and sensitivity
protocol (“mix-and-then-detect”) allows any end users to perform the analy- of the proposed instrumentation in bioassay. The developed system also
sis in their own laboratories. successfully detected a single-particle having larger diameter than the finger
spacing. In addition, the pneumatic pump system was successfully applied
Recently, we studied the classic pentameric lectin, cholera toxin subunit B from delivering the native particles to washing the unbounded particles,
(CTB), binding to GM1-like gangliosides (GM1, fucosyl-GM1, and GM2). We which can enable fast and automated analysis in various bioassays. With
performed experimental measurements and theoretical analysis of a step- these results, we will further improve limit of detection (LOD) by optimizing
wise binding model to investigate the influence of cooperativity on CTB several parameters including the area of the probe surface, a flow rate
binding. Two important phenomena were found. First in contrast to GM1, the during mass transport, hydrodynamic washing step, and the physical proper-
cooperative interaction between bound fucosyl-GM1 molecules is negative. ties of micro/nanoparticles. 79
Surprisingly, such negative binding cooperativity increases the binding
capacity of CTB on a cell membrane surface. We confirmed and explained