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Cincinnati Children's Hospital Medical Center

35 ARTICLES PUBLISHED IN JoVE

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Biology

Single Cell Fate Mapping in Zebrafish
Vikram Kohli 1, Kira Rehn 1, Saulius Sumanas 1,2
1Division of Developmental Biology, Cincinnati Children's Hospital Medical Center, 2Division of Hematology/Oncology, Cincinnati Children's Hospital Medical Center

A method is described to photoactivate single cells containing a caged fluorescent protein using two-photon absorption from a Ti:Sapphire femtosecond laser oscillator. To fate map the photoactivated cell, immunohistochemistry is used. This technique can be applied to any cell type.

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Biology

Slide Preparation Method to Preserve Three-dimensional Chromatin Architecture of Testicular Germ Cells
Satoshi H. Namekawa 1,2
1Division of Reproductive Sciences, Division of Developmental Biology, Perinatal Institute, Cincinnati Children's Hospital Medical Center, 2Department of Pediatrics, University of Cincinnati College of Medicine

The material here describes a method developed to preserve the three-dimensional chromatin structure of testicular germ cells. This has been termed the three-dimensional (3D) slide method. This method improves sensitivity for detection of subnuclear structures and is applicable for immunofluorescence, DNA, and RNA fluorescence in situ hybridization (FISH).

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Biology

Identification of a Murine Erythroblast Subpopulation Enriched in Enucleating Events by Multi-spectral Imaging Flow Cytometry
Diamantis G. Konstantinidis 1, Suvarnamala Pushkaran 1, Katie Giger 1, Stefanos Manganaris 2, Yi Zheng 1, Theodosia A. Kalfa 1
1Cancer and Blood Diseases Institute, Cincinnati Children's Hospital Medical Center, University of Cincinnati College of Medicine, 2IBM

The present protocol describes a novel method of identifying a population of enucleating orthochromatic erythroblasts by multi-spectral imaging flow cytometry, providing a visualization of the erythroblast enucleation process.

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Biology

A Method for Screening and Validation of Resistant Mutations Against Kinase Inhibitors
Meenu Kesarwani 1, Erika Huber 1, Zachary Kincaid 1, Mohammad Azam 1
1Divisions of Experimental Hematology and Cancer Pathology, Cancer Blood Disease Institute, Cincinnati Children's Hospital Medical Center

Emergence of genetic resistance against kinase inhibitor therapy poses significant challenge for effective cancer therapy. Identification and characterization of resistant mutations against a newly developed drug helps in better clinical management and next generation drug design. Here, we describe our protocol for in vitro screening and validation of resistant mutations.

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Biology

Quick Fluorescent In Situ Hybridization Protocol for Xist RNA Combined with Immunofluorescence of Histone Modification in X-chromosome Inactivation
Minghui Yue *1,2, John Lalith Charles Richard *1,2, Norishige Yamada 1,2, Akiyo Ogawa 1, Yuya Ogawa 1,2
1Division of Reproductive Sciences, Cincinnati Children's Hospital Medical Center, 2Department of Pediatrics, University of Cincinnati College of Medicine

We developed an easily customized strand-specific fluorescent in situ hybridization (FISH) protocol combined with immunofluorescence. This allows for a detailed examination of RNA dynamics with simultaneous insight into the chromatin structure, nuclear organization, and transcriptional regulation at the single cell level.

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Medicine

Mouse Pneumonectomy Model of Compensatory Lung Growth
Sheng Liu 1, Jeffrey Cimprich 1, Brian M. Varisco 1
1Division of Critical Care Medicine, Cincinnati Children's Hospital Medical Center

Mouse pneumonectomy is a commonly employed model of compensatory lung growth. This procedure can be used in conjunction with lineage tracing or transgenic mouse models to elucidate underlying mechanisms.

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Medicine

Quantification of the Immunosuppressant Tacrolimus on Dried Blood Spots Using LC-MS/MS
Touraj Shokati 1, Nicholas Bodenberger 1, Holly Gadpaille 1, Björn Schniedewind 1, Alexander A. Vinks 2, Wenlei Jiang 3, Rita R. Alloway 4, Uwe Christians 1
1iC42 Clinical Research and Development, University of Colorado, Anschutz Medical Campus, 2Division of Clinical Pharmacology, Cincinnati Children's Hospital Medical Center, 3Food and Drug Administration (FDA), Center of Drug Evaluation Research - Office of Generic Drugs, 4Transplant Clinical Research, University of Cincinnati

Here we describe a high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) assay to quantify the immunosuppressant tacrolimus in dried blood spots using a simple manual protein precipitation step and online column extraction.

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Medicine

Establishment of Human Epithelial Enteroids and Colonoids from Whole Tissue and Biopsy
Maxime M. Mahe 1, Nambirajan Sundaram 1, Carey L. Watson 1, Noah F. Shroyer 2, Michael A. Helmrath 1
1Department of Pediatric General and Thoracic Surgery, Cincinnati Children's Hospital Medical Center, 2Department of Medicine, Section of Gastroenterology and Hepatology, Baylor College of Medicine

We describe a method to establish human enteroids from small intestinal crypts and colonoids from colon crypts collected from both surgical tissue and biopsies. In this methodological article, we present the culture modalities that are essential for the successful growth and maintenance of human enteroids and colonoids.

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Behavior

Vision Training Methods for Sports Concussion Mitigation and Management
Joseph F. Clark 1, Angelo Colosimo 2, James K. Ellis 3, Robert Mangine 3, Benjamin Bixenmann 4, Kimberly Hasselfeld 2, Patricia Graman 5, Hagar Elgendy 2, Gregory Myer 6, Jon Divine 2
1Neurology and Rehabilitative Medicine, University of Cincinnati, 2Division of Sports Medicine, Department of Orthopaedic Surgery, University of Cincinnati, 3Department of Athletics, University of Cincinnati, 4Department of Neurosurgery, University of Cincinnati, 5College of Education, Criminal Justice, and Human Services, University of Cincinnati, 6Division of Sports Medicine, Cincinnati Children's Hospital Medical Center

This paper describes a protocol to conduct, quantitatively monitor, and assess the success of vision training initiated as part of a sports medical management program including intervention for concussion prevention and performance enhancement.

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Developmental Biology

Using Ex Vivo Upright Droplet Cultures of Whole Fetal Organs to Study Developmental Processes during Mouse Organogenesis
Sarah J. Potter 1, Tony DeFalco 1
1Division of Reproductive Sciences, Cincinnati Children's Hospital Medical Center

The ex vivo upright droplet culture is an alternative to current in vitro and in vivo experimental techniques. This protocol is easy to perform and requires smaller amounts of reagent, while permitting the ability to manipulate and study fetal vascularization, morphogenesis, and organogenesis.

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Medicine

Repeated Measurement of Respiratory Muscle Activity and Ventilation in Mouse Models of Neuromuscular Disease
Victoria N. Jensen *1, Shannon H. Romer *2, Sarah M. Turner 3, Steven A. Crone 3
1Neuroscience Graduate Program, University of Cincinnati, 2Department of Biological Sciences, Wright State University, 3Division of Neurosurgery, Cincinnati Children's Hospital Medical Center

This paper introduces a method for repeated measurements of ventilation and respiratory muscle activity in a freely behaving amyotrophic lateral sclerosis (ALS) mouse model throughout disease progression with whole-body plethysmography and electromyography via an implanted telemetry device.

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Developmental Biology

Histological Analyses of Acute Alcoholic Liver Injury in Zebrafish
Jillian L. Ellis 1, Chunyue Yin 1,2
1Division of Gastroenterology, Hepatology and Nutrition, Cincinnati Children's Hospital Medical Center, 2Division of Developmental Biology, Cincinnati Children's Hospital Medical Center

This protocol describes histological analyses of the livers from zebrafish larvae that have been treated with 2% ethanol for 24 h. Such acute ethanol treatment results in hepatic steatosis and swelling of the hepatic vasculature.

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Chemistry

In Situ Characterization of Boehmite Particles in Water Using Liquid SEM
Juan Yao 1, Bruce W. Arey 1, Li Yang 1, Fei Zhang 1, Rachel Komorek 1, Jaehun Chun 1, Xiao-Ying Yu 1
1Earth & Biological Sciences Directorate, Pacific Northwest National Laboratory

We present a procedure for real-time imaging and elemental composition analysis of boehmite particles in deionized water by in situ liquid Scanning Electron Microscopy.

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Neuroscience

Whole-cell Currents Induced by Puff Application of GABA in Brain Slices
Yangjian Feng *1,2, Binliang Tang *1,2, Ming Chen 2, Li Yang 1,3,4,5
1School of Psychology, South China Normal University, 2School of Life Sciences, South China Normal University, 3Brain Science Institute, South China Normal University, 4Center for Studies of Psychological Application, South China Normal University, 5Guangdong Key Laboratory of Mental Health and Cognitive Science, South China Normal University

We describe the puff technique, by which pharmacological reagents can be administered during whole-cell patch-clamp recording, and highlight various aspects of the features that are crucial for its success.

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Neuroscience

Online Transcranial Magnetic Stimulation Protocol for Measuring Cortical Physiology Associated with Response Inhibition
Michael D. Guthrie 1, Donald L. Gilbert 2, David A. Huddleston 2, Ernest V. Pedapati 2,3, Paul S. Horn 2, Stewart H. Mostofsky 4, Steve W. Wu 2
1College of Medicine, University of Cincinnati, 2Division of Neurology, Cincinnati Children's Hospital Medical Center, 3Division of Psychiatry, Cincinnati Children's Hospital Medical Center, 4Center for Neurodevelopmental and Imaging Research, Kennedy Krieger Institute

We describe an experimental procedure to quantify excitability and inhibition of primary motor cortex during a motor response inhibition task by using Transcranial Magnetic Stimulation throughout the course of a Stop Signal Task.

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Biology

Guided Protocol for Fecal Microbial Characterization by 16S rRNA-Amplicon Sequencing
Ayelet Di Segni 1, Tzipi Braun 1, Marina BenShoshan 1, Sarit Farage Barhom 1, Efrat Glick Saar 1, Karen Cesarkas 1, James E. Squires 2, Nathan Keller 1,3, Yael Haberman 1,4
1Sheba Medical Center, 2Children's Hospital of Pittsburgh of UPMC, 3Ariel University, 4Cincinnati Children's Hospital Medical Center

This manuscript describes a detailed standardized protocol of high-throughput 16S rRNA-amplicon sequencing. The protocol introduces an integrated, uniformed, feasible, and inexpensive protocol starting from fecal sample collection through data analyses. This protocol enables analysis of large numbers of samples with rigorous standards and several controls.

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Biochemistry

A Western Blotting Protocol for Small Numbers of Hematopoietic Stem Cells
Xiongwei Cai 1,2,3, Yi Zheng 1, Nancy A. Speck 2
1Division of Experimental Hematology, Cincinnati Children's Hospital Medical Center, 2Department of Cell and Developmental Biology, Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, 3Department of Obstetrics and Gynecology, Southwest Hospital, Third Military Medical University

A standard Western blotting protocol was optimized for analyzing as few as 500 hematopoietic stem or progenitor cells. Optimization involves careful handling of the cell sample, limiting transfers between tubes, and directly lysing the cells in Laemmli sample buffer.

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Genetics

Production and Purification of Baculovirus for Gene Therapy Application
Md Nasimuzzaman 1,2, Johannes C.M. van der Loo 1,2,3, Punam Malik 1,2
1Division of Experimental Hematology and Cancer Biology, Cincinnati Children's Hospital Medical Center, 2University of Cincinnati College of Medicine, 3Raymond G. Perelman Center for Cellular and Molecular Therapeutics, The Children's Hospital of Philadelphia

In this protocol, baculovirus is produced by transient transfection of baculovirus plasmid into Sf9 cells and amplified in a serum-free suspension culture. The supernatant is purified by heparin affinity chromatography and further concentrated by ultracentrifugation. This protocol is useful for the production and purification of baculovirus for gene therapy application.

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Medicine

Generation of Human Nasal Epithelial Cell Spheroids for Individualized Cystic Fibrosis Transmembrane Conductance Regulator Study
John J. Brewington 1, Erin T. Filbrandt 1, Francis J. LaRosa III 1, Jessica D. Moncivaiz 1, Alicia J. Ostmann 1, Lauren M. Strecker 1, John P. Clancy 1
1Department of Pediatrics, Division of Pulmonary Medicine, Cincinnati Children's Hospital Medical Center

Here we describe a method to generate three-dimensional spheroid cultures of human nasal epithelial cells. Spheroids are then stimulated to drive Cystic Fibrosis Transmembrane Conductance Regulator (CFTR)-dependent ion and fluid secretion, quantifying the change in the spheroid luminal size as a proxy for CFTR function.

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Cancer Research

Methods for Evaluating the Role of c-Fos and Dusp1 in Oncogene Dependence
Meenu Kesarwani 1, Zachary Kincaid 1, Mohammad Azam 1
1Cancer Blood Disease Institute, Divisions of Experimental Hematology and Cancer Pathology, Cincinnati Children's Hospital Medical Center

Here, we describe protocols for the genetic and chemical validation of c-Fos and Dusp1 as a drug target in leukemia using in vitro and in vivo genetic and humanized mouse models. This method can be applied to any target for genetic validation and therapeutic development.

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Medicine

Isolation of Primary Mouse Hepatocytes for Nascent Protein Synthesis Analysis by Non-radioactive L-azidohomoalanine Labeling Method
Esam S.B. Salem *1,2, Kazutoshi Murakami *2, Toshimasa Takahashi 2, Elise Bernhard 2, Vishnupriya Borra 2, Mridula Bethi 2, Takahisa Nakamura 2,3,4
1Department of Pharmacology and Systems Physiology, College of Medicine, University of Cincinnati, 2Division of Endocrinology, Cincinnati Children's Hospital Medical Center, 3Division of Developmental Biology, Cincinnati Children's Hospital Medical Center, 4Department of Pediatrics, College of Medicine, University of Cincinnati

Here, we present a protocol for the isolation of healthy and functional primary mouse hepatocytes. Instructions for detecting hepatic nascent protein synthesis by non-radioactive labeling substrate were provided to help understand the mechanisms underlying protein synthesis in the context of energy-metabolism homeostasis in the liver.

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Bioengineering

Novel Process for 3D Printing Decellularized Matrices
Stacey M. S. Gruber 1, Paulomi Ghosh 2, Karl Wilhelm Mueller 2, Patrick W. Whitlock 1,2,3, Chia-Ying Lin 1,3
1Department of Biomedical Engineering, University of Cincinnati, 2Division of Orthopaedic Surgery, Cincinnati Children's Hospital Medical Center, 3Department of Orthopaedic Surgery, University of Cincinnati

This protocol describes the production of polycaprolactone (PCL) filament with embedded polylactic acid (PLA) microspheres which contain decellularized matrices (DM) for 3D printing of structural tissue engineering constructs.

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Neuroscience

Diagnosis of Hirschsprung's Disease by Immunostaining Rectal Suction Biopsies for Calretinin, S100 Protein and Protein Gene Product 9.5
Shuiqing Chi 1, Mijing Fang 1, Kang Li 1, Li Yang 2, Shao-tao Tang 1
1Department of Pediatric Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, 2Division of Gastroenterology, Hepatology and Nutrition, Cincinnati Children's Hospital Medical Center

This protocol describes the process of immunostaining rectal suction biopsies for calretinin, S100 protein, and protein gene product 9.5. This novel adjuvant diagnostic method for Hirschsprung's disease has preferable sensitivity and specificity rates.

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Immunology and Infection

Co-staining Blood Vessels and Nerve Fibers in Adipose Tissue
Xin Li 1, Zhengmei Mao 2, Li Yang 1, Kai Sun 1
1Center for Metabolic and Degenerative Diseases, the Brown Foundation of Institute of Molecular Medicine, University of Texas Health Science Center at Houston, 2Microscopy Core, the Brown Foundation of Institute of Molecular Medicine, University of Texas Health Science Center at Houston

New blood vessel formation and sympathetic innervation play pivotal roles in adipose tissue remodeling. However, there remain technical issues in visualizing and quantitatively measuring adipose tissue. Here we present a protocol to successfully label and quantitatively compare the densities of blood vessels and nerve fibers in different adipose tissues.

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Neuroscience

Evaluation of Hemisphere Lateralization with Bilateral Local Field Potential Recording in Secondary Motor Cortex of Mice
Yunan Chen 1,2, Ming Li 3, Ying Zheng 3, Li Yang 1
1School of Life Sciences, Guangzhou University, 2Institute for Brain Research and Rehabilitation, South China Normal University, 3School of Life Sciences, South China Normal University

We present in vivo electrophysiological recording of the local field potential (LFP) in bilateral secondary motor cortex (M2) of mice, which can be applied to evaluate hemisphere lateralization. The study revealed altered levels of synchronization between the left and right M2 in APP/PS1 mice compared to WT controls.

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Biology

Purification of Platelets from Mouse Blood
Marilou G. Narciso 1, Md Nasimuzzaman 1,2
1Division of Experimental Hematology and Cancer Biology, Cincinnati Children's Hospital Medical Center, 2University of Cincinnati College of Medicine

Here, mouse blood was collected in the presence of an anti-coagulant. The platelets were purified by iohexol gradient medium using low speed centrifugation. The platelets were activated with thrombin to investigate if they were viable. The quality of the purified platelets was analyzed by flow cytometry and microscopy.

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Developmental Biology

Isolation of Endocardial and Coronary Endothelial Cells from the Ventricular Free Wall of the Rat Heart
Alyssa Klein 1,2,3, Bethel Bayrau 1,2,3, Yifei Miao 1,2,3,4,5, Mingxia Gu 1,2,3,4,5
1Department of Pediatrics, Division of Cardiology, Stanford School of Medicine, 2Vera Moulton Wall Center for Pulmonary Vascular Disease, Stanford School of Medicine, 3Stanford Cardiovascular Institute, Stanford School of Medicine, 4Division of Pulmonary Biology, Department of Pediatrics, Cincinnati Children's Hospital Medical Center, 5Center for Stem Cell and Organoid Medicine, CuSTOM, Division of Developmental Biology, and Perinatal Institute, Cincinnati Children's Hospital Medical Center

We present a protocol for the isolation of endocardial and coronary endothelial cells from rat hearts through sequential tissue digestion in a digestion buffer, cell collection from recurrent centrifuge cycles, and cell purification using anti-rat CD31 microbeads.

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Developmental Biology

A Patient-Derived Xenograft Model for Venous Malformation
Sandra Schrenk *1, Jillian Goines *1, Elisa Boscolo 1,2
1Division of Experimental Hematology and Cancer Biology, Cincinnati Children's Hospital Medical Center, 2Department of Pediatrics, University of Cincinnati College of Medicine

We present a detailed protocol to generate a murine xenograft model of venous malformation. This model is based on the subcutaneous injection of patient-derived endothelial cells containing hyper-activating TIE2 and/or PIK3CA gene mutations. Xenograft lesions closely recapitulate the histopathological features of VM patient tissue.

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Developmental Biology

Isolation of Murine Spermatogenic Cells using a Violet-Excited Cell-Permeable DNA Binding Dye
Yu-Han Yeh *1,2, Mengwen Hu *1,2, Toshinori Nakagawa 3,4, Akihiko Sakashita 1,2, Shosei Yoshida 3,4, So Maezawa 5,6, Satoshi H. Namekawa 1,2
1Division of Reproductive Sciences, Division of Developmental Biology, Perinatal Institute, Cincinnati Children's Hospital Medical Center, 2Department of Pediatrics, University of Cincinnati College of Medicine, 3Division of Germ Cell Biology, National Institute for Basic Biology, National Institutes of Natural Sciences, 4Department of Basic Biology, School of Life Science, Graduate University for Advanced Studies (Sokendai), 5Department of Animal Science and Biotechnology, School of Veterinary Medicine, Azabu University, 6Faculty of Science and Technology, Department of Applied Biological Science, Tokyo University of Science

Here we present a simple and efficient method to isolate live meiotic and post-meiotic germ cells from adult mouse testes. Using a low-cytotoxicity, violet-excited DNA binding dye and fluorescence-activated cell sorting, one can isolate highly enriched spermatogenic cell populations for many downstream applications.

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Medicine

Refined CLARITY-Based Tissue Clearing for Three-Dimensional Fibroblast Organization in Healthy and Injured Mouse Hearts
Demetria M. Fischesser 1,2, Evan C. Meyer 3, Michelle Sargent 2, Jeffery D. Molkentin 2
1Department of Molecular Genetics, Biochemistry, and Microbiology, University of Cincinnati College of Medicine, 2Division of Molecular Cardiovascular Biology, Cincinnati Children's Hospital Medical Center, 3Confocal Imaging Core, Cincinnati Children's Hospital Medical Center

A refined method of tissue clearing was developed and applied to the adult mouse heart. This method was designed to clear dense, autofluorescent cardiac tissue, while maintaining labeled fibroblast fluorescence attributed to a genetic reporter strategy.

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Biology

Process Development for the Production and Purification of Adeno-Associated Virus (AAV)2 Vector using Baculovirus-Insect Cell Culture System
Md Nasimuzzaman 1,2, Sophia Villaveces 1, Johannes C. M. van der Loo 1,2,3, Sivani Alla 1
1Division of Experimental Hematology and Cancer Biology, Cincinnati Children's Hospital Medical Center, 2University of Cincinnati College of Medicine, 3Raymond G. Perelman Center for Cellular and Molecular Therapeutics, The Children's Hospital of Philadelphia

In this protocol, AAV2 vector is produced by co-culturing Spodoptera frugiperda (Sf9) insect cells with baculovirus (BV)-AAV2-green fluorescent protein (GFP) or therapeutic gene and BV-AAV2-rep-cap infected Sf9 cells in suspension culture. AAV particles are released from the cells using detergent, clarified, purified by affinity column chromatography, and concentrated by tangential flow filtration.

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Bioengineering

Combining Human Organoids and Organ-on-a-Chip Technology to Model Intestinal Region-Specific Functionality
Gauri Kulkarni *1, Athanasia Apostolou *1, Lorna Ewart 1, Carolina Lucchesi 1, Magdalena Kasendra 2,3
1Emulate Inc. Boston, 2Center for Stem Cell and Organoid Medicine, Cincinnati Children’s Hospital Medical Center, 3Division of Pediatric General and Thoracic Surgery, Cincinnati Children’s Hospital Medical Center

Biopsy-derived intestinal organoids and organ-on-a-chips technologies are combined into a microphysiological platform to recapitulate region-specific intestinal functionality.

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Biology

An Integrated Approach for Microprotein Identification and Sequence Analysis
Omar Brito-Estrada *1, Keira R. Hassel *1, Catherine A. Makarewich 1,2
1The Heart Institute, Division of Molecular Cardiovascular Biology, Cincinnati Children's Hospital Medical Center, 2Department of Pediatrics, University of Cincinnati College of Medicine

The protocol described here provides detailed instructions on how to analyze genomic regions of interest for microprotein-coding potential using PhyloCSF on the user-friendly UCSC Genome Browser. Additionally, several tools and resources are recommended to further investigate sequence characteristics of identified microproteins to gain insight into their putative functions.

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Bioengineering

Reconstituting Cytoarchitecture and Function of Human Epithelial Tissues on an Open-Top Organ-Chip
Varone Antonio 1, Adya Panchal 2, Magdalena Kasendra 3, Barrile Riccardo 2,3
1Merk Sharp & Dohme LLC, 2Department of Biomedical Engineering, University of Cincinnati, 3Center for Stem Cell and Organoid Medicine, Cincinnati Children's Hospital Medical Center

The present protocol describes the capabilities and the essential culture modalities of the Open-Top Organ-Chip for the successful establishment and maturation of full-thickness organ-on-chip cultures of primary tissues (skin, alveolus, airway, and intestine), providing the opportunity to investigate different functional aspects of the human epithelial/mesenchymal and vascular niche interface in vitro.

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Bioengineering

Hyperpolarized 129Xe Lung MRI and Spectroscopy in Mechanically Ventilated Mice
Mariah L. Costa 1,2,3, Joseph W. Plummer 1,2, Abdullah S. Bdaiwi 1, Brice J. Albert 4, Elizabeth M. Fugate 3, Peter J. Niedbalski 5,6,7, Diana M. Lindquist 3,8, Zackary I. Cleveland 1,2,3,8,9
1Center for Pulmonary Imaging Research, Division of Pulmonary Medicine, Cincinnati Children's Hospital Medical Center, 2Department of Biomedical Engineering, University of Cincinnati, 3Imaging Research Center, Department of Radiology, Cincinnati Children's Hospital Medical Center, 4Analytical Technology Group, Aurorium, 5Division of Pulmonary, Critical Care, and Sleep Medicine, Department of Internal Medicine, University of Kansas Medical Center, 6Department of Bioengineering, University of Kansas, 7Hoglund Biomedical Imaging Center, University of Kansas Medical Center, 8Department of Pediatrics, University of Cincinnati, 9Division of Pulmonary Medicine, Cincinnati Children's Hospital Medical Center

Hyperpolarized xenon MRI can quantify regional lung microstructure (air-space dimensions) and physiology (ventilation and gas exchange) in translational research and clinical care. Although challenging, it can provide comparable pulmonary insights in preclinical studies. This protocol describes the infrastructure and procedures needed to perform routine xenon lung MRI in mice.

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