University of Illinois, Urbana-Champaign View Institution's Website 65 articles published in JoVE Cancer Research Application of AlDeSense to Stratify Ovarian Cancer Cells Based on Aldehyde Dehydrogenase 1A1 Activity Michael C. Lee*1,3,4, Sarah H. Gardner*2,3,4, Rodrigo Tapia Hernandez1,3,4, Jefferson Chan1,2,3,4 1Department of Chemistry, University of Illinois Urbana-Champaign, 2Department of Biochemistry, University of Illinois Urbana-Champaign, 3Beckman Institute for Advanced Science and Technology, University of Illinois Urbana-Champaign, 4Cancer Center of Illinois, University of Illinois Urbana-Champaign Methods to measure ALDH1A1 activity in live cells are critical in cancer research due to its status as a biomarker of stemness. In this study, we employed an isoform-selective fluorogenic probe to determine the relative levels of ALDH1A1 activity in a panel of five ovarian cancer cell lines. Biology Isolation and Characterization of the Murine Uterosacral Ligaments and Pelvic Floor Organs Catalina S. Bastías1, Lea M. Savard2, Kevin N. Eckstein2, Kathleen Connell3, Callan M. Luetkemeyer2,5, Virginia L. Ferguson1,2,4, Sarah Calve1,2,4 1Biomedical Engineering Program, University of Colorado Boulder, 2Paul M. Rady Department of Mechanical Engineering, University of Colorado Boulder, 3Department of Obstetrics and Gynecology, University of Colorado Anschutz Medical Campus, 4BioFrontiers Institute, University of Colorado Boulder, 5Department of Mechanical Science and Engineering, University of Illinois Urbana-Champaign This article presents a detailed protocol for dissecting uterosacral ligaments and other pelvic floor tissues, including the cervix, rectum, and bladder in mice, to expand the study of female reproductive tissues. Engineering Performing Microscope-Mounted Y-Shaped Cutting Tests Matthew Guerena*1, Jing-Chen Peng*1, Marcus Schmid*1, Cecilia Walsh*1, Shaobo Zhan*1, Shelby B. Hutchens1 1Mechanical Science and Engineering, University of Illinois at Urbana-Champaign Y-shaped cutting measures fracture-relevant length scales and energies in soft materials. Previous apparatuses were designed for benchtop measurements. This protocol describes the fabrication and use of an apparatus that orients the setup horizontally and provides the fine positioning capabilities necessary for in situ viewing, plus failure quantification, via an optical microscope. Bioengineering In Vitro Selection of Aptamers to Differentiate Infectious from Non-Infectious Viruses Marcos Ezequiel Gramajo*1, Ryan J. Lake*2, Yi Lu3, Ana Sol Peinetti1 1INQUIMAE (CONICET), Departamento de Química Inorgánica, Analítica y Química Física, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, 2Department of Chemistry, University of Illinois at Urbana-Champaign, 3Department of Chemistry, University of Texas at Austin We provide a protocol that can be generally applied to select aptamers that bind to infectious viruses only and not to viruses that have been rendered non-infectious by a disinfection method or to any other similar viruses. This opens the possibility of determining infectivity status in portable and rapid tests. Biochemistry Single-Molecule Diffusion and Assembly on Polymer-Crowded Lipid Membranes Satyaghosh Maurya1, Vishwesh Haricharan Rai1, Aditya Upasani1, Saurabh Umrao1,2, Diksha Parwana1, Rahul Roy1,3 1Department of Chemical Engineering, Indian Institute of Science, 2Holonyak Micro & Nanotechnology Lab, University of Illinois, Urbana-Champaign, 3Center for BioSystems Science and Engineering, Indian Institute of Science Here, a protocol to perform and analyze the binding, mobility, and assembly of single molecules on artificial crowded lipid membranes using single-molecule total internal reflection fluorescence (smTIRF) microscopy is presented. Biology High-Throughput Analysis of Non-Photochemical Quenching in Crops Using Pulse Amplitude Modulated Chlorophyll Fluorometry Dhananjay Gotarkar1,2, Lynn Doran1,2, Meghan Burns1,2, Abigail Hinkle1, Johannes Kromdijk1,3, Steven J. Burgess1,2 1Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, 2Department of Plant Biology, Morrill Hall, University of Illinois at Urbana-Champaign, 3Environmental Plant Physiology group, Department of Plant Sciences, University of Cambridge The protocol introduces a high-throughput method for measuring the relaxation of non-photochemical quenching by pulse amplitude modulated chlorophyll fluorometry. The method is applied to field-grown Glycine max and can be adapted to other species to screen for genetic diversity or breeding populations. Chemistry Characterizing RNA Modifications in Single Neurons Using Mass Spectrometry Kevin D. Clark1,2, Stanislav S. Rubakhin1,2, Jonathan V. Sweedler1,2 1Beckman Institute for Advanced Science and Technology, University of Illinois Urbana-Champaign, 2Department of Chemistry, University of Illinois Urbana-Champaign Post-transcriptional modifications of RNA represent an understudied layer of translation regulation that has recently been linked to central nervous system plasticity. Here, sample preparation and liquid chromatography-tandem mass spectrometry approach is described for simultaneous characterization of numerous RNA modifications in single neurons. Biochemistry Use of Microscale Thermophoresis to Measure Protein-Lipid Interactions Robert P. Sparks*1, William Lawless*2, Andres S. Arango*1,3,4, Emad Tajkhorshid1,3,4, Rutilio A. Fratti1,3 1Department of Biochemistry, University of Illinois at Urbana-Champaign, 2Department of Chemistry, University of South Florida, 3Center for Biophysics and Quantitative Biology, University of Illinois at Urbana-Champaign, 4Beckman Institute for Advanced Science and Technology, University of Illinois at Urbana-Champaign Microscale thermophoresis obtains binding constants quickly at low material cost. Either labeled or label free microscale thermophoresis is commercially available; however, label free thermophoresis is not capable of the diversity of interaction measurements that can be performed using fluorescent labels. We provide a protocol for labeled thermophoresis measurements. Engineering Assembly and Characterization of an External Driver for the Generation of Sub-Kilohertz Oscillatory Flow in Microchannels Giridar Vishwanathan1, Gabriel Juarez1 1Department of Mechanical Science and Engineering, University of Illinois at Urbana-Champaign The protocol demonstrates a convenient method to produce harmonic oscillatory flow from 10-1000 Hz in microchannels. This is performed by interfacing a computer-controlled speaker diaphragm to the microchannel in a modular manner. Neuroscience Extraction and Dissection of the Domesticated Pig Brain Stephen A. Fleming1, Supida Monaikul2, Austin T. Mudd1, Reeba Jacob3, Ryan N. Dilger1,2,3 1Neuroscience Program, University of Illinois at Urbana-Champaign, 2Department of Animal Sciences, University of Illinois at Urbana-Champaign, 3Division of Nutritional Sciences, University of Illinois at Urbana-Champaign This protocol details the technique for removal of the pig brain in its entirety and dissection of several brain regions commonly studied in neuroscience. Medicine Psychophysical Tracking Method to Assess Taste Detection Thresholds in Children, Adolescents, and Adults: The Taste Detection Threshold (TDT) Test Paule V. Joseph1, Julie A. Mennella2, Beverly J. Cowart2, M. Yanina Pepino3 1National Institute of Alcohol Abuse and Alcoholism, Section of Sensory Science and Metabolism, 2Monell Chemical Senses Center, 3Department of Food Science and Human Nutrition, University of Illinois at Urbana Champaign Psychophysical tools measure the functionality of the taste system for both research and health assessment purposes. This paper describes a method to measure taste detection thresholds that can determine the lowest concentration of sucrose, sodium chloride, or monosodium glutamate that can be tasted by individuals as young as 6 years. Biology Assessing Agrochemical Risk to Mated Honey Bee Queens Julia D. Fine1, Kendall M. Torres2, Jamilyn Martin2, Gene E. Robinson2,3,4 1Invasive Species and Pollinator Health Research Unit, USDA-ARS, 2Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, 3Neuroscience Program, University of Illinois at Urbana-Champaign, 4Department of Entomology, University of Illinois at Urbana-Champaign This protocol was developed to enhance the understanding of how agrochemicals affect honey bee (Apis mellifera) reproduction by establishing methods to expose honey bee queens and their worker caretakers to agrochemicals in a controlled, laboratory setting and carefully monitoring their relevant responses. Biology Preparation of Virus-Enriched Inoculum for Oral Infection of Honey Bees (Apis mellifera) Edward M. Hsieh1, Jimena Carrillo-Tripp2, Adam G. Dolezal1 1Department of Entomology, University of Illinois, 2Department of Microbiology, Center for Scientific Research and Higher Education of Ensenada (CICESE) Here we describe two protocols: first to propagate, extract, purify, and quantify large quantities of honey bee non-enveloped virus particles, including a method for removing honey bee pupae and second to test the effects of viral infection using a highly repeatable, high-throughput cage bioassay. Biology Probing mRNA Kinetics in Space and Time in Escherichia coli using Two-Color Single-Molecule Fluorescence In Situ Hybridization Sangjin Kim1,2,3,4, Kavya Vaidya1,2 1Department of Physics, University of Illinois at Urbana-Champaign, 2Center for the Physics of Living Cells, University of Illinois at Urbana-Champaign, 3Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, 4Center for Biophysics and Quantitative Biology, University of Illinois at Urbana-Champaign This protocol describes an application of single-molecule fluorescence in situ hybridization (smFISH) to measure the in vivo kinetics of mRNA synthesis and degradation. Developmental Biology Imaging Intranuclear Actin Rods in Live Heat Stressed Drosophila Embryos Natalie Biel1,2, Lauren Figard3, Anna Marie Sokac1,2,3 1Integrative Molecular and Biomedical Sciences, Baylor College of Medicine, 2Department of Cell and Molecular Biology, School of Molecular and Cellular Biology, University of Illinois at Urbana-Champaign, 3Verna and Marrs McLean Department of Biochemistry and Molecular Biology, Baylor College of Medicine The goal of this protocol is to inject Rhodamine-conjugated globular actin into Drosophila embryos and image intranuclear actin rod assembly following heat stress. Engineering Studying Large Amplitude Oscillatory Shear Response of Soft Materials Johnny Ching-Wei Lee1, Jun Dong Park1, Simon A. Rogers1 1Department of Chemical and Biomolecular Engineering, University of Illinois at Urbana-Champaign We present a detailed protocol outlining how to perform nonlinear oscillatory shear rheology on soft materials, and how to run the SPP-LAOS analysis to understand the responses as a sequence of physical processes. Cancer Research Capturing Small Molecule Communication Between Tissues and Cells Using Imaging Mass Spectrometry Katherine E. Zink1, Matthew Dean1,2, Joanna E. Burdette1, Laura M. Sanchez1 1Department of Medicinal Chemistry and Pharmacognosy, University of Illinois at Chicago, 2Department of Animal Science, University of Illinois at Urbana-Champaign A novel method of sample preparation was developed to accommodate cell and tissue coculture to detect small molecule exchange using imaging mass spectrometry. Medicine Protocol and Guidelines for Point-of-Care Lung Ultrasound in Diagnosing Neonatal Pulmonary Diseases Based on International Expert Consensus Jing Liu1,2, Roberto Copetti3, Erich Sorantin4, Jovan Lovrenski5, Javier Rodriguez-Fanjul6, Dalibor Kurepa7, Xing Feng8, Luigi Cattaross9, Huayan Zhang10,11, Misun Hwang12, Tsu F. Yeh13,14, Yisrael Lipener7, Abhay Lodha15, Jia-Qin Wang16, Hai-Ying Cao2,17, Cai-Bao Hu2,18, Guo-Rong Lyu19, Xin-Ru Qiu1,2, Li-Qun Jia20, Xiao-Man Wang20, Xiao-Ling Ren1,2, Jiu-Ye Guo1,2, Yue-Qiao Gao1,2, Jian-Jun Li1,2, Ying Liu1,2, Wei Fu1,2, Yan Wang21, Zu-Lin Lu1,2, Hua-Wei Wang8, Li-Li Shang22 1Department of Neonatology and NICU, Beijing Chaoyang District Maternal and Child Healthcare Hospital, 2The Neonatal Lung Ultrasound Training Base, Chinese College of Critical Ultrasound, 3Emergency Department, Cattinara University Hospital, 4Division of Pediatric Radiology, Department of Radiology, Medical University Graz, 5Faculty of Medicine, University of Novi Sad, Radiology Department, Institute for Children and Adolescents Health Care of Vojvodina, 6Pediatric Intensive Care Unit, Pediatric Service Hospital Joan XXIII Tarragona, University Rovira i Virgil, 7Department of Neonatology, Children's Hospital of Soochow University, 9Department of Neonatology, Udine University Hospital, 10Division of Neonatology, Children's Hospital of Philadelphia, 12Intensive Care Unit, Zhejiang Hospital, 19Collaborative Innovation Center for Maternal and Infant Health Service Application Technology, Quanzhou Medical College, 20Department of Neonatology and NICU, Tai'an City Central Hospital of Shandong Province, 22Department of Intensive Care Unit, The Second Affiliated Hospital of Heilongjiang University of Chinese Medicine Lung ultrasound is a noninvasive and valuable tool for bedside evaluation of neonatal lung diseases. However, a relative lack of reference standards, protocols and guidelines may limit its application. Here, we aim to develop a standardized neonatal lung ultrasound diagnostic protocol to be used in clinical decision-making. Medicine Cloud-Based Phrase Mining and Analysis of User-Defined Phrase-Category Association in Biomedical Publications Dibakar Sigdel*1,2, Vincent Kyi*1,2, Aiden Zhang*1, Shaun P. Setty3, David A. Liem1,2,4, Yu Shi5, Xuan Wang5, Jiaming Shen5, Wei Wang1,6,7, JiaWei Han5, Peipei Ping1,2,4,6 1The NIH BD2K Center of Excellence in Biomedical Computing, University of California, Los Angeles, 2Department of Physiology, University of California, Los Angeles, 3Department of Pediatric and Adult Congenital Heart Surgery, Miller Children's and Women's Hospital and Long Beach Memorial Hospital, 4Department of Medicine/Cardiology, University of California, Los Angeles, 5NIH BD2K Program Centers of Excellence for Big Data Computing -- KnowEng Center, Department of Computer Science, University of Illinois at Urbana-Champaign (UIUC), 6Scalable Analytics Institute (ScAi), University of California, Los Angeles, 7Department of Computer Science, University of California, Los Angeles We present a protocol and associated programming code as well as metadata samples to support a cloud-based automated identification of phrases-category association representing unique concepts in user selected knowledge domain in biomedical literature. The phrase-category association quantified by this protocol can facilitate in depth analysis in the selected knowledge domain. Environment Improving Infrared Spectroscopy Characterization of Soil Organic Matter with Spectral Subtractions Andrew J. Margenot1, Sanjai J. Parikh2, Francisco J. Calderón3 1Department of Crop Sciences, University of Illinois Urbana-Champaign, 2Department of Land, Air and Water Resources, University of California Davis, 3Central Great Plains Resources Management Research, USDA ARS SOM underlies many soil functions and processes, but its characterization by FTIR spectroscopy is often challenged by mineral interferences. The described method can increase the utility of SOM analysis by FTIR spectroscopy by subtracting mineral interferences in soil spectra using empirically obtained mineral reference spectra. Behavior Probing the Limits of Egg Recognition Using Egg Rejection Experiments Along Phenotypic Gradients Lindsay Canniff1, Miri Dainson2, Analía V. López3, Mark E. Hauber2, Tomáš Grim4, Peter Samaš5, Daniel Hanley1 1Department of Biology, Long Island University-Post, 2Department of Animal Biology, School of Integrative Biology, University of Illinois, 3Departamento de Ecología, Genética y Evolución, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, 4Department of Zoology and Laboratory of Ornithology, Palacký University, 5Institute of Vertebrate Biology, Academy of Sciences of the Czech Republic This protocol provides guidelines for running egg rejection experiments: outlining techniques for painting experimental egg models to emulate the colors of natural bird eggs, conducting fieldwork, and analyzing the collected data. This protocol provides a uniform method for conducting comparable egg rejection experiments. Immunology and Infection Using Phylogenetic Analysis to Investigate Eukaryotic Gene Origin Dechun Zhang*1, Xianzhao Kan*2, Sarah Elizabeth Huss3, Lan Jiang2, Li-Qing Chen3, Yibing Hu4 1Key Laboratory of Three Gorges Regional Plant Genetics and Germplasm Enhancement (CTGU)/Biotechnology Research Center, China Three Gorges University, 2The Institute of Bioinformatics, College of Life Sciences, Anhui Normal University, 3Department of Plant Biology, University of Illinois at Urbana-Champaign, 4College of Resources & Environmental Sciences, Nanjing Agricultural University A method of constructing a phylogenetic tree based on sequence homology of SWEETs from eukaryotes and SemiSWEETs from prokaryotes is described. Phylogenetic analysis is a useful tool for explaining the evolutionary relatedness between homologous proteins or genes from different organism groups. Environment High-throughput, Microscale Protocol for the Analysis of Processing Parameters and Nutritional Qualities in Maize (Zea mays L.) Carrie Butts-Wilmsmeyer1, Nicole A. Yana1, Gurshagan Kandhola2, Kent D. Rausch2, Rita H. Mumm1, Martin O. Bohn1 1Department of Crop Sciences, University of Illinois at Urbana-Champaign, 2Department of Agricultural and Biological Engineering, University of Illinois at Urbana-Champaign Here, we present a microscale protocol for processing grain samples and for incorporating this microscale approach into a high-throughput analytical pipeline. This is a higher throughput adaptation of currently available protocols. Bioengineering Fragility Assessment of Bovine Cortical Bone Using Scratch Tests Kavya Mendu1, Amrita Kataruka1, Jasmine Puthuvelil1, Ange-Therese Akono1,2 1Department of Civil and Environmental Engineering, University of Illinois at Urbana Champaign, 2Department of Mechanical Science and Engineering, University of Illinois at Urbana Champaign This study assesses the fracture toughness of bovine cortical bone at the sub-meso levels using microscopic scratch tests. This is an original, objective, rigorous, and reproducible method proposed to probe fracture toughness below the macroscopic scale. Potential applications are studying changes in bone fragility due to diseases like osteoporosis. Bioengineering A Method to Estimate Cadaveric Femur Cortical Strains During Fracture Testing Using Digital Image Correlation Timothy Rossman1, Susheil Uthamaraj1, Asghar Rezaei1,2, Sean McEligot1, Hugo Giambini3, Iwona Jasiuk4, Michael J. Yaszemski3, Lichun Lu3, Dan Dragomir-Daescu1,2 1Division of Engineering, Mayo Clinic, 2Department of Physiology and Biomedical Engineering, Mayo Clinic, 3Department of Orthopedic Surgery, Mayo Clinic, 4Department of Mechanical Science and Engineering, University of Illinois at Urbana-Champaign In this protocol, the femur surface strains are estimated during fracture testing using the digital image correlation technique. The novelty of the method involves application of a high-contrast stochastic speckle pattern on the femur surface, carefully specified illumination, high speed video capture, and digital image correlation analysis for strain calculations. Bioengineering Method and Instrumented Fixture for Femoral Fracture Testing in a Sideways Fall-on-the-Hip Position Dan Dragomir-Daescu1,2, Asghar Rezaei1,2, Timothy Rossman2, Susheil Uthamaraj2, Rachel Entwistle2, Sean McEligot2, Vincent Lambert2, Hugo Giambini3, Iwona Jasiuk4, Michael J. Yaszemski3, Lichun Lu1,3 1Department of Physiology and Biomedical Engineering, Mayo Clinic, 2Division of Engineering, Mayo Clinic, 3Department of Orthopedic Surgery, Mayo Clinic, 4Department of Mechanical Science and Engineering, University of Illinois at Urbana-Champaign In this manuscript, we present a protocol to fracture test cadaveric proximal femora in a sideways fall on the hip configuration using instrumented fixtures mounted on a standard servo hydraulic frame. Nine digitized signals comprising forces, moments, and displacement along with two high speed video streams are acquired during testing. Neuroscience Electroconvulsive Seizures in Rats and Fractionation of Their Hippocampi to Examine Seizure-induced Changes in Postsynaptic Density Proteins Sung-Soo Jang1,2, Han Gil Jeong1, Hee Jung Chung1,2 1Department of Molecular and Integrative Physiology, University of Illinois at Urbana-Champaign, 2Neuroscience Program, University of Illinois at Urbana-Champaign Electroconvulsive seizure (ECS) is an experimental animal model of electroconvulsive therapy for severe depression. ECS globally stimulates activity in the hippocampus, leading to synaptogenesis and synaptic plasticity. Here, we describe methods for ECS induction in rats and for subcellular fractionation of their hippocampi to examine seizure-induced changes in synaptic proteins. Developmental Biology Light-mediated Reversible Modulation of the Mitogen-activated Protein Kinase Pathway during Cell Differentiation and Xenopus Embryonic Development Vishnu V. Krishnamurthy*1, Aurora J. Turgeon*2, John S. Khamo1, Payel Mondal1, Savanna R. Sharum1, Wenyan Mei2, Jing Yang2, Kai Zhang1,3,4 1Department of Biochemistry, University of Illinois at Urbana-Champaign, 2Department of Comparative Biosciences, University of Illinois at Urbana-Champaign, 3Neuroscience Program, University of Illinois at Urbana-Champaign, 4Center for Biophysics and Quantitative Biology, University of Illinois at Urbana-Champaign This protocol describes an optogenetic strategy to modulate mitogen-activated protein kinase (MAPK) activity during cell differentiation and Xenopus embryonic development. This method allows for the reversible activation of the MAPK signaling pathway in mammalian cell culture and in multicellular live organisms, like Xenopus embryos, with high spatial and temporal resolution. Medicine Proximal Cadaveric Femur Preparation for Fracture Strength Testing and Quantitative CT-based Finite Element Analysis Dan Dragomir-Daescu1,2, Asghar Rezaei1,2, Susheil Uthamaraj2, Timothy Rossman2, James T. Bronk3, Mark Bolander3, Vincent Lambert2, Sean McEligot2, Rachel Entwistle2, Hugo Giambini3, Iwona Jasiuk4, Michael J. Yaszemski3, Lichun Lu1,3 1Department of Physiology and Biomedical Engineering, Mayo Clinic, 2Division of Engineering, Mayo Clinic, 3Department of Orthopedic Surgery, Mayo Clinic, 4Department of Mechanical Science and Engineering, University of Illinois at Urbana-Champaign We present a robust protocol on how to carefully preserve and prepare cadaveric femora for fracture testing and quantitative computed tomography imaging. The method provides precise control over input conditions for the purpose of determining relationships between bone mineral density, fracture strength, and defining finite element model geometry and properties. Bioengineering A High-throughput Cell Microarray Platform for Correlative Analysis of Cell Differentiation and Traction Forces Kerim B. Kaylan1, Andreas P. Kourouklis1, Gregory H. Underhill1 1Department of Bioengineering, University of Illinois at Urbana–Champaign Cell differentiation is regulated by a host of microenvironmental factors, including both matrix composition and substrate material properties. We describe here a technique utilizing cell microarrays in conjunction with traction force microscopy to evaluate both cell differentiation and biomechanical cell–substrate interactions as a function of microenvironmental context. Bioengineering Merging Ion Concentration Polarization between Juxtaposed Ion Exchange Membranes to Block the Propagation of the Polarization Zone Minyoung Kim1,2, Hyunjoon Rhee1,3, Ji Yoon Kang1, Tae Song Kim1, Rhokyun Kwak1 1Center for BioMicrosystems, Korea Institute of Science and Technology, 2Department of Mechanical Engineering, Seoul National University, 3Department of Industrial Engineering, University of Illinois Urbana-Champaign The protocol for a novel ion concentration polarization (ICP) platform that can stop the propagation of the ICP zone, regardless of the operating conditions is described. This unique ability of the platform lies in the use of merging ion depletion and enrichment, which are two polarities of the ICP phenomenon. Cancer Research A Method of Targeted Cell Isolation via Glass Surface Functionalization Ali Ansari1, Reema Patel2, Kinsey Schultheis1, Vesna Naumovski3, P. I. Imoukhuede1 1Department of Bioengineering, University of Illinois at Urbana-Champaign, 2Department of Liberal Arts & Sciences, University of Illinois at Urbana-Champaign, 3Department of Biomedical Engineering, Illinois Institute of Technology This protocol describes customizable surface functionalization of the desthiobiotin, streptavidin, and APTES system in order to isolate specific cell types of interest. In addition, this manuscript covers the applications, optimization, and verification of this process. Bioengineering Fabrication of a Functionalized Magnetic Bacterial Nanocellulose with Iron Oxide Nanoparticles Sandra L. Arias1, Akshath R. Shetty2, Angana Senpan3, Mónica Echeverry-Rendón4, Lisa M. Reece5,6, Jean Paul Allain1,2,3,7 1Department of Bioengineering, University of Illinois at Urbana-Champaign, 2Department of Nuclear, Plasma and Radiological Engineering, University of Illinois at Urbana-Champaign, 3Micro and Nanotechnology Laboratory, University of Illinois at Urbana-Champaign, 4Program of Study and Control of Tropical Diseases (PECET), University of Antioquia, 5Sealy Center for Vaccine Development, University of Texas Medical Branch, 6WHO Collaborating Center for Vaccine Research, Evaluation and Training on Emerging Infectious Diseases, University of Texas Medical Branch, 7Beckman Institute, University of Illinois at Urbana-Champaign Here, we present a protocol to make a bacterial nanocellulose (BNC) magnetic for applications in damaged blood vessel reconstruction. The BNC was synthesized by G. xylinus strain. On the other hand, magnetization of the BNC was realized through in situ precipitation of Fe2+ and Fe3+ ferrous ions inside the BNC mesh. Medicine Systems Biology of Metabolic Regulation by Estrogen Receptor Signaling in Breast Cancer Yiru Chen Zhao1, Zeynep Madak Erdogan1 1Department of Food Sciences and Human Nutrition, University of Illinois, Urbana-Champaign Integration of data from genome-wide sequencing experiments and metabolomics experiments is a challenge. In this paper we report, for the first time, generation, analysis and integration of transcriptome, cistrome and metabolome data from breast cancer cells treated with estradiol. Engineering Three-dimensional Particle Tracking Velocimetry for Turbulence Applications: Case of a Jet Flow Jin-Tae Kim1, David Kim1, Alex Liberzon2, Leonardo P. Chamorro1,3 1Department of Mechanical Science and Engineering, University of Illinois at Urbana-Champaign, 2School of Mechanical Engineering, Tel Aviv University, 3Department of Civil and Environmental Engineering, University of Illinois at Urbana-Champaign A three-dimensional particle tracking velocimetry (3D-PTV) system based on a high-speed camera with a four-view splitter is described here. The technique is applied to a jet flow from a circular pipe in the vicinity of ten diameters downstream at Reynolds number Re ≈ 7,000. Medicine Rapid Fractionation and Isolation of Whole Blood Components in Samples Obtained from a Community-based Setting Amy Weckle1,2, Allison E. Aiello3, Monica Uddin1,4, Sandro Galea5, Rebecca M. Coulborn6, Richelo Soliven7, Helen Meier6, Derek E. Wildman1,2 1Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, 2Department of Molecular and Integrative Physiology, University of Illinois at Urbana-Champaign, 3Department of Epidemiology, Gillings School of Global Public Health, University of North Carolina, 4Department of Psychology, University of Illinois at Urbana-Champaign, 5Department of Epidemiology, Mailman School of Public Health, Columbia University, 6Department of Epidemiology, University of Michigan School of Public Health, 7Center for Molecular Medicine and Genetics, Wayne State University School of Medicine We outline a methodology for the processing of whole blood to obtain a variety of components for further analysis. We have optimized a streamlined protocol that enables rapid, high-throughput simultaneous processing of whole blood samples in a non-clinical setting. Neuroscience Modification of a Colliculo-thalamocortical Mouse Brain Slice, Incorporating 3-D printing of Chamber Components and Multi-scale Optical Imaging Bernard J. Slater1, Anthony Y. Fan2, Kevin A. Stebbings1, M. Taher A. Saif2, Daniel A. Llano1,3 1Neuroscience Program, University of Illinois Urbana-Champaign, 2Mechanical Science and Engineering, University of Illinois Urbana-Champaign, 3Molecular and Integrative Physiology, University of Illinois Urbana-Champaign Using multiple angles to cut the mouse pup brain, we improve upon a previously-described acute brain slice which captures the connections between most of the major auditory midbrain and forebrain structures. Environment Measuring Rates of Herbicide Metabolism in Dicot Weeds with an Excised Leaf Assay Rong Ma1, Joshua J. Skelton1, Dean E. Riechers1 1Department of Crop Sciences, University of Illinois This manuscript describes how herbicide metabolism rates can be effectively quantified with excised leaves from a dicot weed, thereby reducing variability and removing any possible confounding effects of herbicide uptake or translocation typically observed in whole-plant assays. Medicine A Hydrogel Construct and Fibrin-based Glue Approach to Deliver Therapeutics in a Murine Myocardial Infarction Model. Molly Melhem1, Tor Jensen1, Larissa Reinkensmeyer1, Luke Knapp1, Jordan Flewellyn1, Lawrence Schook1 1Department of Bioengineering, University of Illinois at Urbana-Champaign This protocol aims to alleviate the limitation of poor cell engraftment for stem cell treatment of myocardial infarctions through the use of a hydrogel system and a fibrin-based glue. With this approach, cell-to-tissue contact post-infarction can be maintained, increasing the therapeutic potential of beneficial agents at the site of injury. Bioengineering Isolation of Primary Human Colon Tumor Cells from Surgical Tissues and Culturing Them Directly on Soft Elastic Substrates for Traction Cytometry M. Yakut Ali1, Sandeep V. Anand1, Krishnarao Tangella2,3, Davendra Ramkumar2,3, Taher A. Saif1,4 1Department of Mechanical Science and Engineering, University of Illinois at Urbana-Champaign, 2College of Medicine, University of Illinois at Urbana-Champaign, 3Provena Covenant Medical Centre, 4Micro and Nanotechnology Laboratory, University of Illinois at Urbana-Champaign A protocol is described to extract primary human cells from surgical colon tumor and normal tissues. The isolated cells are then cultured on soft elastic substrates (polyacrylamide hydrogels) functionalized by an extracellular matrix protein, and embedded with fluorescent microbeads. Traction cytometry is performed to assess cellular contractile stresses. Biology Fluorescence Imaging with One-nanometer Accuracy (FIONA) Yong Wang*1,2, En Cai*1,2, Janet Sheung1,2, Sang Hak Lee1,2, Kai Wen Teng2,3, Paul R. Selvin1,2,3 1Department of Physics, University of Illinois at Urbana-Champaign, 2Center for the Physics of Living Cells, University of Illinois at Urbana-Champaign, 3Center for Biophysics and Computational Biology, University of Illinois at Urbana-Champaign Single fluorophores can be localized with nanometer precision using FIONA. Here a summary of the FIONA technique is reported, and how to carry out FIONA experiments is described. Bioengineering A Novel Method for Localizing Reporter Fluorescent Beads Near the Cell Culture Surface for Traction Force Microscopy Samantha G. Knoll1, M. Yakut Ali1, M. Taher A. Saif1 1Department of Mechanical Science and Engineering, University of Illinois at Urbana-Champaign Traditional techniques for fabricating polyacrylamide (PA) gels containing fluorescent probes involve sandwiching a gel between an adherent surface and a glass slide. Here, we show that coating this slide with poly-D-lysine (PDL) and fluorescent probes localizes the probes to within 1.6 µm from the gel surface. Environment Multimodal Optical Microscopy Methods Reveal Polyp Tissue Morphology and Structure in Caribbean Reef Building Corals Mayandi Sivaguru1, Glenn A. Fried1, Carly A. H. Miller1,2, Bruce W. Fouke1,2,3 1Institute for Genomic Biology, University of Illinois at Urbana-Champaign, 2Department of Geology, University of Illinois at Urbana-Champaign, 3Department of Microbiology, University of Illinois at Urbana-Champaign An integrated suite of imaging techniques has been applied to determine polyp morphology and tissue structure in the Caribbean corals Montastraeaannularis and M. faveolata. Fluorescence, serial block face, and two-photon confocal laser scanning microscopy have identified lobate structure, polyp walls, and estimated chromatophore and zooxanthellae densities and distributions. Neuroscience In Vivo Imaging of Dauer-specific Neuronal Remodeling in C. elegans Nathan E. Schroeder1, Kristen M. Flatt1 1Department of Crop Sciences, University of Illinois Urbana-Champaign Following exposure to specific environmental stressors, the nematode Caenorhabditis elegans undergoes extensive phenotypic plasticity to enter into a stress-resistant ‘dauer’ juvenile stage. We present methods for the controlled induction and imaging of neuroplasticity during dauer. Engineering Micro-masonry for 3D Additive Micromanufacturing Hohyun Keum1, Seok Kim1 1Mechanical Science and Engineering, University of Illinois at Urbana-Champaign This paper introduces a 3D additive micromanufacturing strategy (termed ‘micro-masonry’) for the flexible fabrication of microelectromechanical system (MEMS) structures and devices. This approach involves transfer printing-based assembly of micro/nanoscale materials in conjunction with rapid thermal annealing-enabled material bonding techniques. Neuroscience A Comprehensive Protocol for Manual Segmentation of the Medial Temporal Lobe Structures Matthew Moore*1, Yifan Hu*1, Sarah Woo1, Dylan O'Hearn1, Alexandru D. Iordan2,3, Sanda Dolcos1, Florin Dolcos1,2,3 1Psychology Department, University of Illinois Urbana-Champaign, 2Neuroscience Program, University of Illinois Urbana-Champaign, 3Beckman Institute for Advanced Science and Technology, University of Illinois Urbana-Champaign The present work provides a comprehensive set of guidelines for manually tracing the medial temporal lobe (MTL) structures. This protocol can be applied to research involving structural and/or combined structural-functional magnetic resonance imaging (MRI) investigations of the MTL, in both healthy and clinical groups. Engineering Fabrication and Testing of Microfluidic Optomechanical Oscillators Kewen Han1, Kyu Hyun Kim2, Junhwan Kim1, Wonsuk Lee2,3, Jing Liu3, Xudong Fan3, Tal Carmon2, Gaurav Bahl1 1Mechanical Science and Engineering, University of Illinois at Urbana-Champaign, 2Electrical Engineering and Computer Science, University of Michigan, 3Biomedical Engineering, University of Michigan Parametric optomechanical excitations have recently been experimentally demonstrated in microfluidic optomechanical resonators by means of optical radiation pressure and stimulated Brillouin scattering. This paper describes the fabrication of these microfluidic resonators along with methodologies for generating and verifying optomechanical oscillations. Biology Visualizing Cytoplasmic Flow During Single-cell Wound Healing in Stentor coeruleus Mark Slabodnick1,2, Bram Prevo1,3, Peter Gross1,4, Janet Sheung1,5, Wallace Marshall1,2 1Physiology Course, Marine Biological Laboratory, 2Department of Biochemistry & Biophysics, University of California San Francisco, 3Department of Physics and Astronomy, Vrije Universiteit Amsterdam, 4Max Planck Institute of Molecular Cell Biology and Genetics, 5Department of Physics, University of Illinois Urbana-Champaign The giant ciliate Stentor coeruleus is a classical system for studying regeneration and wound healing in single cells. By imaging Stentor cells simultaneously at low and high magnification it is possible to measure cytoplasmic flows before, during, and after wounding. Behavior Mouse Short- and Long-term Locomotor Activity Analyzed by Video Tracking Software Jason M. York1, Neil A. Blevins2, Leslie K. McNeil2, Gregory G. Freund1,2 1Department of Animal Sciences, University of Illinois at Urbana-Champaign, 2Department of Pathology, Integrative Immunology and Behavior Program, University of Illinois at Urbana-Champaign Locomotor activity (LMA) is a simple and easily performed measurement of behavior in mice. Coupling of video tracking software (VTS) and LMA allows for the improvement of specificity and sensitivity, especially when compared with the manual, line crossing method of LMA analysis. Additionally VTS allows long-term tracking of mouse LMA. Bioengineering Micropipette Aspiration of Substrate-attached Cells to Estimate Cell Stiffness Myung-Jin Oh1, Frank Kuhr1, Fitzroy Byfield2, Irena Levitan1 1Section of Respiratory, Critical Care and Sleep Medicine, Department of Medicine, University of Illinois, 2Institute for Medicine and Engineering, University of Pennsylvania Here we describe a quick and simple method to measure cell stiffness. The general principle of this approach is to measure membrane deformation in response to well-defined negative pressure applied through a micropipette to the cell surface. This method provides a powerful tool to study biomechanical properties of substrate-attached cells. Neuroscience Brain Imaging Investigation of the Impairing Effect of Emotion on Cognition Gloria Wong1,2, Sanda Dolcos1,3, Ekaterina Denkova1, Rajendra Morey4,5,6, Lihong Wang4,5, Gregory McCarthy6,7, Florin Dolcos1,2,3,8,9 1Department of Psychiatry, University of Alberta, 2Centre for Neuroscience, University of Alberta, 3Department of Psychology, University of Illinois, 4Brain Imaging and Analysis Center, Duke University, 5Department of Psychiatry and Behavioral Sciences, Duke University, 6Mid-Atlantic Mental Illness Research Education and Clinical Center, VA Medical Center, 7Department of Psychology, Yale University, 8Neuroscience Program, University of Illinois, 9Beckman Institute for Advanced Science & Technology, University of Illinois We present a protocol that allows investigation of the neural mechanisms mediating the detrimental impact of emotion on cognition, using functional magnetic resonance imaging. This protocol can be used with both healthy and clinical participants. Biology Two Types of Assays for Detecting Frog Sperm Chemoattraction Lindsey A. Burnett1, Nathan Tholl2, Douglas E. Chandler2 1Department of Animal Sciences, University of Illinois, Urbana-Champaign, 2School of Life Sciences, Arizona State University Eggs and the extracellular coatings around eggs frequently release peptides, proteins and small molecules that communicate with sperm to guide them to the egg thereby promoting fertilization. Using frog sperm we describe and compare two classes of assays used to detect sperm chemoattraction – sperm accumulation assays and sperm tracking assays. Bioengineering Specimen Preparation, Imaging, and Analysis Protocols for Knife-edge Scanning Microscopy Yoonsuck Choe1, David Mayerich2, Jaerock Kwon3, Daniel E. Miller1, Chul Sung1, Ji Ryang Chung1, Todd Huffman4, John Keyser1, Louise C. Abbott5 1Department of Computer Science and Engineering, Texas A&M University, 2Beckman Institute for Advanced Science and Technology, University of Illinois, 3Department of Electrical and Computer Engineering, Kettering University, 43Scan, 5Department of Veterinary Integrative Biosciences, Texas A&M University The full process from brain specimen preparation to serial sectioning imaging using the Knife-Edge Scanning Microscope, to data visualization and analysis is described. This technique is currently used to acquire mouse brain data, but it is applicable to other organs, other species. Bioengineering Planar and Three-Dimensional Printing of Conductive Inks Bok Yeop Ahn1, Steven B. Walker1, Scott C. Slimmer1, Analisa Russo1, Ashley Gupta1, Steve Kranz1, Eric B. Duoss1,2, Thomas F. Malkowski1,3, Jennifer A. Lewis1 1Department of Materials Science and Engineering, University of Illinois at Urbana-Champaign, 2Center for Micro- and Nanotechnology, Lawrence Livermore National Laboratory, 3Presently at the Interdisciplinary Center for Wide Band-gap Semiconductors, University Of California Santa Barbara Planar and three-dimensional printing of conductive metallic inks is described. Our approach provides new avenues for fabricating printed electronic, optoelectronic, and biomedical devices in unusual layouts at the microscale. Immunology and Infection Following Cell-fate in E. coli After Infection by Phage Lambda Lanying Zeng1, Ido Golding1,2,3 1Department of Physics, University of Illinois at Urbana-Champaign, 2Center for the Physics of Living Cells, University of Illinois at Urbana-Champaign, 3Verna and Marrs McLean Department of Biochemistry and Molecular Biology, Baylor College of Medicine This article describes the procedure for preparing a fluorescently-labeled version of bacteriophage lambda, infection of E. coli bacteria, following the infection outcome under the microscope, and analysis of infection results. Neuroscience Brain Imaging Investigation of the Neural Correlates of Emotion Regulation Sanda Dolcos1, Keen Sung2, Ekaterina Denkova3, Roger A. Dixon4,5, Florin Dolcos1,6,7 1Department of Psychology, University of Illinois, Urbana-Champaign, 2Department of Computing Science, University of Alberta, Edmonton, 3Department of Psychiatry, University of Alberta, Edmonton, 4Department of Psychology, University of Alberta, Edmonton, 5Centre for Neuroscience, University of Alberta, Edmonton, 6Neuroscience Program, University of Illinois, Urbana-Champaign, 7Beckman Institute, University of Illinois, Urbana-Champaign We present a protocol that allows investigation of the neural correlates of deliberate and automatic emotion regulation, using functional magnetic resonance imaging. This protocol can be used in healthy participants, both young and older, as well as in clinical patients. Neuroscience Brain Imaging Investigation of the Neural Correlates of Emotional Autobiographical Recollection Ekaterina Denkova1, Trisha Chakrabarty1, Sanda Dolcos1,2, Florin Dolcos1,2,3,4 1Department of Psychiatry, University of Alberta, Edmonton, 2Psychology Department, University of Illinois, Urbana-Champaign, 3Neuroscience Program, University of Illinois, Urbana-Champaign, 4Beckman Institute for Advanced Science & Technology, University of Illinois, Urbana-Champaign We present a protocol that allows investigation of the neural correlates of recollecting emotional autobiographical memories, using functional magnetic resonance imaging. This protocol can be used with both healthy and clinical participants. Neuroscience Brain Imaging Investigation of the Neural Correlates of Observing Virtual Social Interactions Keen Sung1, Sanda Dolcos2, Sophie Flor-Henry3, Crystal Zhou3, Claudia Gasior4, Jennifer Argo5, Florin Dolcos2,6,7 1Department of Computing Science, University of Alberta, 2Department of Psychology, University of Illinois, 3Centre for Neuroscience, University of Alberta, 4Department of Psychology, University of Alberta, 5Department of Marketing, Business Economics, and Law, University of Alberta, 6Neuroscience Program, University of Illinois at Urbana-Champaign, 7Beckman Institute, University of Illinois at Urbana-Champaign This article demonstrates an experimental design in which whole-body animated characters are used in conjunction with functional magnetic resonance imaging (fMRI) to investigate the neural correlates of observing virtual social interactions. Neuroscience Brain Imaging Investigation of the Memory-Enhancing Effect of Emotion Andrea Shafer1, Alexandru Iordan2, Roberto Cabeza3, Florin Dolcos1,4 1Centre for Neuroscience, University of Alberta, 2Neuroscience Program, University of Illinois, Urbana-Champaign, 3Center for Cognitive Neuroscience, Duke University, 4Psychology Department, Neuroscience Program, & Beckman Institute, University of Illinois, Urbana-Champaign We present a protocol that uses functional magnetic resonance imaging to investigate the neural correlates of the memory-enhancing effect of emotion. This protocol allows identification of brain activity specifically linked to memory-related processing, contrary to more general perceptual processing, and can be used with healthy and clinical populations. Bioengineering Separating Beads and Cells in Multi-channel Microfluidic Devices Using Dielectrophoresis and Laminar Flow Larry J. Millet1,2, Kidong Park1,2, Nicholas N. Watkins1,2, K. Jimmy Hsia2,3, Rashid Bashir1,2,4 1Electrical and Computer Engineering, University of Illinois at Urbana-Champaign, 2Micro and Nanotechnology Lab, University of Illinois at Urbana-Champaign, 3Mechanical Science and Engineering, University of Illinois at Urbana-Champaign, 4Bioengineering, University of Illinois at Urbana-Champaign Dielectrophoresis (DEP) is an effective method to manipulate cells. Printed circuit boards (PCB) can provide inexpensive, reusable and effective electrodes for contact-free cell manipulation within microfluidic devices. By combining PDMS-based microfluidic channels with coverslips on PCBs, we demonstrate bead and cell manipulation and separation within multichannel microfluidic devices. Bioengineering A Microfluidic-based Hydrodynamic Trap for Single Particles Eric M. Johnson-Chavarria1, Melikhan Tanyeri2, Charles M. Schroeder1,2 1Center for Biophysics and Computational Biology, University of Illinois at Urbana-Champaign, 2Department of Chemical and Biomolecular Engineering, University of Illinois at Urbana-Champaign In this article, we present a microfluidic-based method for particle confinement based on hydrodynamic flow. We demonstrate stable particle trapping at a fluid stagnation point using a feedback control mechanism, thereby enabling confinement and micromanipulation of arbitrary particles in an integrated microdevice. Biology Rapid Homogeneous Detection of Biological Assays Using Magnetic Modulation Biosensing System Amos Danielli1,2, Noga Porat3, Marcelo Ehrlich4, Ady Arie1 1Department of Physical Electronics, Faculty of Engineering, Tel Aviv University, 2Department of Biomedical Engineering, Washington University in St. Louis, 3Department of Biological Sciences, University of Illinois, 4Department of Cell Research and Immunology, Tel Aviv University Magnetic modulation biosensing system is utilized to rapidly, sensitively and simply detect biological assays, such as DNA molecules and proteins. Biology Fabrication and Operation of an Oxygen Insert for Adherent Cellular Cultures Shawn Oppegard1, Elly Sinkala1, David Eddington1 1Bioengineering, University of Illinois Fabrication and validation of an add-on platform that offers enhanced control over the spatial and temporal oxygenation in a 6-well plate. The device is adaptable to a number of culture systems and can be used to investigate the effects of oxygen on wound healing. Biology Electrophysiological Recording in the Drosophila Embryo Kaiyun Chen1, David E. Featherstone1, Kendal Broadie2 1Department of Biological Sciences, University of Illinois, 2Department of Biological Sciences, Vanderbilt University Electrophysiological recordings from Drosophila embryos allow analyses of developing muscle and neuron electrical properties, as well as characterization of functional synaptogenesis at the glutamatergic neuromuscular junction and central cholinergic and GABAergic synapses. Biology Harvesting and Preparing Drosophila Embryos for Electrophysiological Recording and Other Procedures David E. Featherstone1, Kaiyun Chen1, Kendal Broadie2 1Department of Biological Sciences, University of Illinois, 2Department of Biological Sciences, Vanderbilt University This technique exposes the Drosophila embryonic neuromusculature for immunohistochemistry or electrophysiological recording. It is useful for studying early events in neuromuscular development or performing electrophysiology in mutants that cannot hatch.