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Memorial Sloan Kettering Cancer Center

27 ARTICLES PUBLISHED IN JoVE

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Biology

Visualization of the Interstitial Cells of Cajal (ICC) Network in Mice
Yu Chen 1,2, Tambudzai Shamu 2, Hui Chen 3, Peter Besmer 3, Charles L. Sawyers 2,4, Ping Chi 1,5
1Department of Medicine, Memorial Sloan Kettering Cancer Center, 2Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, 3Developmental Biology Program, Memorial Sloan Kettering Cancer Center, 4Howard Hughes, Medical Institute, 5Laboratory of Chromatin Biology and Epigenetics, The Rockefeller University

The interstitial cells of Cajal (ICC) are the pacemaker cells of the gastrointestinal (GI) tract. They form complex networks between smooth muscle cells and post-ganglionic neuronal fibers to regulate GI contractility. Here, we present immunofluorescence methods cross-sectional and whole-mount visualization of murine ICC networks.

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Medicine

A Laser-induced Mouse Model of Chronic Ocular Hypertension to Characterize Visual Defects
Liang Feng 1,2, Hui Chen 1, Genn Suyeoka 1, Xiaorong Liu 1,2
1Department of Ophthalmology, Northwestern University, 2Department of Neurobiology, Northwestern University

Chronic ocular hypertension is induced using laser photocoagulation of the trabecular meshwork in mouse eyes. The intraocular pressure (IOP) is elevated for several months after laser treatment. The decrease of visual acuity and contrast sensitivity of experimental animals are monitored using the optomotor test.

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Chemistry

Microfluidic On-chip Capture-cycloaddition Reaction to Reversibly Immobilize Small Molecules or Multi-component Structures for Biosensor Applications
Carlos Tassa 1, Monty Liong 1, Scott Hilderbrand 1, Jason E. Sandler 1, Thomas Reiner 1, Edmund J. Keliher 1, Ralph Weissleder 1, Stanley Y. Shaw 1
1Center for Systems Biology, Massachusetts General Hospital

We present a method for rapid, reversible immobilization of small molecules and functionalized nanoparticle assemblies for Surface Plasmon Resonance (SPR) studies, using sequential on-chip bioorthogonal cycloaddition chemistry and antibody-antigen capture.

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Medicine

Ex Vivo Treatment Response of Primary Tumors and/or Associated Metastases for Preclinical and Clinical Development of Therapeutics
Adriana D. Corben *1, Mohammad M. Uddin *2, Brooke Crawford 3, Mohammad Farooq 4, Shanu Modi 5, John Gerecitano 5, Gabriela Chiosis 2, Mary L. Alpaugh 6
1Department of Pathology, Memorial Sloan Kettering Cancer Center, 2Molecular Pharmacology and Chemistry Program, Memorial Sloan Kettering Cancer Center, 3Department of Radiology, Weill Cornell Medical College, 4Department of Medicine, Memorial Sloan Kettering Cancer Center, 5Department of Oncology, Memorial Sloan Kettering Cancer Center, 6Department of Surgery, Memorial Sloan Kettering Cancer Center

Established cancer cell lines and xenografts have been the mainstay of cancer research for the past several decades. However, recent evidence suggests that therapeutic response is greatly influenced by the tumor cell microenvironment. Therefore, we have developed an ex vivo analysis of primary tumor specimens for drug development purposes.

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Bioengineering

Harnessing the Bioorthogonal Inverse Electron Demand Diels-Alder Cycloaddition for Pretargeted PET Imaging
Thomas Reiner *1, Jason S. Lewis 1, Brian M. Zeglis *1
1Department of Radiology, Memorial Sloan Kettering Cancer Center

The bioorthogonal inverse electron demand Diels-Alder cycloaddition has been harnessed to create an effective and modular pretargeted PET imaging strategy for cancer. In this protocol, the steps of this methodology are described in the context of a model system employing the colorectal cancer targeted antibody huA33 and a 64Cu-labeled radioligand.

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JoVE Core

The Bioconjugation and Radiosynthesis of 89Zr-DFO-labeled Antibodies
Brian M. Zeglis 1, Jason S. Lewis 1
1Department of Radiology, Memorial Sloan Kettering Cancer Center

Due to its multi-day radioactive half-life and favorable decay properties, the positron-emitting radiometal 89Zr is extremely well-suited for use in antibody-based radiopharmaceuticals for PET imaging. In this protocol, the bioconjugation, radiosynthesis, and preclinical application of 89Zr-labeled antibodies will be described.

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Biology

TRAP-rc, Translating Ribosome Affinity Purification from Rare Cell Populations of Drosophila Embryos
Benjamin Bertin 1, Yoan Renaud 1, Rajaguru Aradhya 2, Krzysztof Jagla 1, Guillaume Junion 1
1University of Clermont-Ferrand, 2Memorial Sloan Kettering Cancer Center

Translating Ribosome Affinity Purification (TRAP) is able to capture cell-type-specific translation of mRNA. Here we report the first TRAP protocol dedicated to isolation of mRNA in rare cell populations of Drosophila embryos.

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Medicine

Studying the Role of Alveolar Macrophages in Breast Cancer Metastasis
Surya Kumari Vadrevu 1, Sharad Sharma 1,2, Navin Chintala 1,3, Jalpa Patel 1, Magdalena Karbowniczek 1, Maciej Markiewski 1
1Department of Immunotherapeutics and Biotechnology, School of Pharmacy, Texas Tech University Health Science Center, 2Merck Research Labs, 3Department of Surgery, Memorial Sloan Kettering Cancer Center

Here we describe the model and approach to study functions of pulmonary alveolar macrophages in cancer metastasis. To demonstrate the role of these cells in metastasis, the syngeneic (4T1) model of breast cancer in conjunction with the depletion of alveolar macrophage with clodronate liposomes was used.

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Immunology and Infection

Rapid Molecular Detection and Differentiation of Influenza Viruses A and B
Caitlin C. Otto 1, Samuel E. Kaplan 1, Jeffrey Stiles 1, Albina Mikhlina 1, Cindy Lee 1, N. Esther Babady 1, Yi-Wei Tang 1
1Department of Laboratory Medicine, Memorial Sloan Kettering Cancer Center

We describe a rapid, molecular-based Influenza A and B assay. The Influenza assay detects each target within 15 min by employing isothermal amplification with influenza-specific primers followed by target detection with molecular beacon probes. The Influenza A and B assay is user-friendly and required minimal hands-on time to perform.

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Genetics

Flow-sorting and Exome Sequencing of the Reed-Sternberg Cells of Classical Hodgkin Lymphoma
Jonathan B. Reichel 1, Jason McCormick 2, Jonathan R. Fromm 3, Olivier Elemento 4, Ethel Cesarman 5, Mikhail Roshal 6
1Innovation Laboratory, Center for Molecular Oncology, Memorial Sloan Kettering Cancer Center, 2Flow-Sorting Core Facility, Weill Cornell Medical College, 3Department of Laboratory Medicine, University of Washington, 4Department of Physiology and Biophysics, Institute for Computational Biomedicine, Weill Cornell Medical College, 5Department of Pathology and Laboratory Medicine, Weill Cornell Medical College, 6Department of Laboratory Medicine, Memorial Sloan Kettering Cancer Center

Here, we describe a combined flow cytometric cell sorting and low-input, next-generation library construction protocol designed to produce high-quality, whole-exome data from the Hodgkin Reed-Sternberg (HRS) cells of classical Hodgkin lymphoma (CHL).

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Bioengineering

A Novel Technique for Generating and Observing Chemiluminescence in a Biological Setting
Gabriel E. Büchel 1,2, Brandon Carney 1,3, Jun Tang 1, Brian M. Zeglis 1,3, Jörg Eppinger 2, Thomas Reiner 1,4
1Department of Radiology, Memorial Sloan Kettering Cancer Center, 2KAUST Catalysis Center, King Abdullah University of Science and Technology, 3Department of Chemistry, Hunter College, and PhD Program in Chemistry, Graduate Center of City University of New York, 4Department of Radiology, Weill Cornell Medical College

This protocol describes a new intraoperative imaging technique that uses a ruthenium complex as a source of chemiluminescent light emission, thereby producing high signal-to-noise ratios during in vivo imaging. Intraoperative imaging is an expanding field that could revolutionize the way that surgical procedures are performed.

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Cancer Research

A Comprehensive Procedure to Evaluate the In Vivo Performance of Cancer Nanomedicines
Jun Tang 1, Carlos Pérez-Medina 1,2, Yiming Zhao 2, Ahmad Sadique 1, Willem J. M. Mulder 2, Thomas Reiner 1
1Department of Radiology, Memorial Sloan Kettering Cancer Center, 2Translational and Molecular Imaging Institute, Icahn School of Medicine at Mount Sinai

The poor understanding of the in vivo performance of nanomedicines stymies their clinical translation. Procedures to evaluate the in vivo behavior of cancer nanomedicines at systemic, tissue, single-cell, and subcellular levels in tumor-bearing immunocompetent mice are described here. This approach may help researchers to identify promising cancer nanomedicines for clinical translation.

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Behavior

Using Laser Doppler Imaging and Monitoring to Analyze Spinal Cord Microcirculation in Rat
Yingli Jing *1,2,3,4, Fan Bai *1,2,3,4, Hui Chen 1,2,3,4, Hao Dong 1,2,3,4
1China Rehabilitation Research Center, 2Institute of Rehabilitation Science of China, 3Center of Neural Injury and Repair, Beijing Institute for Brain Disorders, 4Beijing Key Laboratory of Neural Injury and Rehabilitation

Here we present a combination of laser Doppler perfusion imaging (LDPI) and laser Doppler perfusion monitoring (LDPM) to measure spinal cord local blood flows and oxygen saturation (SO2), as well as a standardized procedure for introducing spinal cord trauma on rat.

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Cancer Research

An In Vivo Murine Sciatic Nerve Model of Perineural Invasion
Sylvie Deborde *1, Yasong Yu *1, Andrea Marcadis 1, Chun-Hao Chen 1, Ning Fan 2, Richard L. Bakst 3, Richard J. Wong 1
1Department of Surgery, Memorial Sloan Kettering Cancer Center, 2Molecular Cytology Core Facility, Memorial Sloan Kettering Cancer Center, 3Department of Radiation Oncology, Mount Sinai Hospital

We describe an in vivo murine model of perineural invasion by injecting syngeneic pancreatic cancer cells into the sciatic nerve. The model allows for quantification of the extent of nerve invasion, and supports investigation of the cellular and molecular mechanisms of perineural invasion.

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Cancer Research

Surface-enhanced Resonance Raman Scattering Nanoprobe Ratiometry for Detecting Microscopic Ovarian Cancer via Folate Receptor Targeting
Chrysafis Andreou 1, Anton Oseledchyk 1, Fay Nicolson 1, Naxhije Berisha 1,2, Suchetan Pal 1, Moritz F. Kircher 1,3,4,5,6,7
1Department of Radiology, Memorial Sloan Kettering Cancer Center, 2Department of Chemistry, The Graduate Center of the City University of New York, 3Molecular Pharmacology Program, Memorial Sloan Kettering Cancer Center, 4Center for Molecular Imaging and Nanotechnology (CMINT), Memorial Sloan Kettering Cancer Center, 5Gerstner Sloan Kettering Graduate School of Biomedical Sciences, 6Department of Radiology, Weill Cornell Medical College of Cornell University, 7Dana-Farber Cancer Institute and Harvard Medical Center

Ovarian cancer forms metastases throughout the peritoneal cavity. Here, we present a protocol to make and use folate-receptor targeted surface-enhanced resonance Raman scattering nanoprobes that reveal these lesions with high specificity via ratiometric imaging. The nanoprobes are administered intraperitoneally to living mice, and the derived images correlate well with histology.

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Chemistry

Pretargeted Radioimmunotherapy Based on the Inverse Electron Demand Diels-Alder Reaction
Rosemery Membreno 1,2, Brendon E. Cook 1,2,3, Brian M. Zeglis 1,2,3,4
1Department of Chemistry, Hunter College of the City University of New York, 2Ph.D. Program in Chemistry, Graduate Center of the City University of New York, 3Department of Radiology, Memorial Sloan Kettering Cancer Center, 4Department of Radiology, Weill Cornell Medical College

This protocol describes the synthesis and characterization of a trans-cyclooctene (TCO)-modified antibody and a 177Lu-labeled tetrazine (Tz) radioligand for pretargeted radioimmunotherapy (PRIT). In addition, it details the use of these two constructs for in vivo biodistribution and longitudinal therapy studies in a murine model of colorectal cancer.

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Chemistry

Synthesis and Bioconjugation of Thiol-Reactive Reagents for the Creation of Site-Selectively Modified Immunoconjugates
Maria Davydova 1, Guillaume Dewaele Le Roi 1,2, Pierre Adumeau 1, Brian M. Zeglis 1,2,3,4
1Department of Chemistry, Hunter College of the City University of New York, 2Ph.D. Program in Chemistry, Graduate Center of the City University of New York, 3Department of Radiology, Memorial Sloan Kettering Cancer Center, 4Department of Radiology, Weill Cornell Medical College

In this protocol, we will describe the synthesis of PODS, a phenyoxadiazolyl methyl sulfone-based reagent for the site-selective attachment of cargos to the thiols of biomolecules, particularly antibodies. In addition, we will describe the synthesis and characterization of a PODS-bearing bifunctional chelator and its conjugation to a model antibody.

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Cancer Research

Prostate Organoid Cultures as Tools to Translate Genotypes and Mutational Profiles to Pharmacological Responses
Kyrie J. Pappas 1, Danielle Choi 1, Charles L. Sawyers 1,2, Wouter R. Karthaus 1
1Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, 2Howard Hughes Medical Institute

Presented here is a protocol to study pharmacological responses in prostate epithelial organoids. Organoids closely resemble in vivo biology and recapitulate patient genetics, making them attractive model systems. Prostate organoids can be established from wildtype prostates, genetically engineered mouse models, benign human tissue, and advanced prostate cancer.

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Cancer Research

In Vitro Establishment of a Genetically Engineered Murine Head and Neck Cancer Cell Line using an Adeno-Associated Virus-Cas9 System
Manu Prasad *1,2, Sankar Jagadeeshan *1,2, Maurizio Scaltriti 3, Irit Allon 2,4, Moshe Elkabets 1,2
1The Shraga Segal Department of Microbiology, Immunology and Genetics, Ben-Gurion University of the Negev, 2Faculty of Health Sciences, Ben-Gurion University of the Negev, 3Human Oncology & Pathogenesis Program (HOPP), Memorial Sloan Kettering Cancer Center, 4Institute of Pathology, Barzilai University Medical Center

Development of murine models with specific genes mutated in head and neck cancer patients is required for understanding of neoplasia. Here, we present a protocol for in vitro transformation of primary murine tongue cells using an adeno-associated virus-Cas9 system to generate murine HNC cell lines with specific genomic alterations.

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JoVE Core

Fabrication of Compressed Hosiery and Measurement of its Pressure Characteristic Exerted on the Lower Limbs
Guangwu Sun 1,2,3, Jiecong Li 4, Xiaona Chen 1,2,3, Yanmei Li 1,2,3, Yu Chen 1, Qi Fang 1, Hong Xie 1,2,3
1School of Fashion Engineering, Shanghai University of Engineering Science, 2Textile Industrial Key Laboratory of Ergonomics and Functional Clothing, Shanghai University of Engineering Science, 3Sino-British Joint Lab for Smart Sportswear, Shanghai University of Engineering Science, 4College of Textiles, Donghua University

This article reports fabrication, structure and pressure measurement of compressed hosiery by employing direct and indirect methods.

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Biochemistry

An In Vitro Single-Molecule Imaging Assay for the Analysis of Cap-Dependent Translation Kinetics
Anthony Gaba *1, Hongyun Wang *1, Xiaohui Qu 1
1Molecular Biology Program, Memorial Sloan Kettering Cancer Center

Tracking individual translation events allows for high-resolution kinetic studies of cap-dependent translation mechanisms. Here we demonstrate an in vitro single-molecule assay based on imaging interactions between fluorescently labeled antibodies and epitope-tagged nascent peptides. This method enables single-molecule characterization of initiation and peptide elongation kinetics during active in vitro cap-dependent translation.

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Cancer Research

Characterization and Functional Prediction of Bacteria in Ovarian Tissues
Lanbo Zhao *1, Weichu Zhao *2, Qi Wang *1, Dongxin Liang 1, Yu Liu 3, Guoxing Fu 4, Lu Han 1, Yiran Wang 1, Chao Sun 1, Qing Wang 1, Qing Song 1, Qiling Li 1, Qinrui Lu 1
1Department of Obstetrics and Gynecology, First Affiliated Hospital of Xi’an Jiaotong University, 2School of Social Sciences, University of Manchester, 3Department of Pathology, First Affiliated Hospital of Xi’an Jiaotong University, 4Omega Bioservices Inc

Immunohistochemistry staining and 16S ribosomal RNA gene (16S rRNA gene) sequencing were performed in order to discover and distinguish bacteria in cancerous and noncancerous ovarian tissues in situ. The compositional and functional differences of the bacteria were predicted by using BugBase and Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt).

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Bioengineering

Viscoelastic Characterization of Soft Tissue-Mimicking Gelatin Phantoms using Indentation and Magnetic Resonance Elastography
Yuan Feng 1, Suhao Qiu 1, Yu Chen 1, Runke Wang 1, Zhao He 1, Linghan Kong 1, Yi Chen 1, Shengyuan Ma 1
1School of Biomedical Engineering, Shanghai Jiao Tong University

This article presents a demonstration and summary of protocols of making gelatin phantoms that mimic soft tissues, and the corresponding viscoelastic characterization using indentation and magnetic resonance elastography.

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Cancer Research

Combining Reflectance Confocal Microscopy with Optical Coherence Tomography for Noninvasive Diagnosis of Skin Cancers via Image Acquisition
Ucalene Harris 1, Milind Rajadhyaksha 1, Manu Jain 1
1Dermatology Service, Department of Medicine, Memorial Sloan Kettering Cancer Center

Here, we describe protocols for acquiring good-quality images using novel, noninvasive imaging devices of reflectance confocal microscopy (RCM) and combined RCM and optical coherence tomography (OCT). We also familiarize clinicians with their clinical applications so that they can integrate the techniques into regular clinical workflows to improve patient care.

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Medicine

Augmented Reality Navigation-Guided Core Decompression for Osteonecrosis of Femoral Head
Qi Wang *1,3, Qiuyuan Wang *2,3, Ran Ding 3, Youjie Yao 4, Junjun Pan 5, Weiguo Wang 3
1China-Japan Friendship School of Clinical Medicine, Peking University, 2Graduate School of Beijing University of Chinese Medicine, 3Department of Orthopedic Surgery, Beijing Key Lab of Immunes-Mediated Inflammatory Disease, China-Japan Friendship Hospital, 4Beijing Normal University, 5State Key Laboratory of Virtual Reality Technology and Systems, Beihang University

Augmented reality technology was applied to core decompression for osteonecrosis of the femoral head to realize real-time visualization of this surgical procedure. This method can effectively improve the safety and precision of core decompression.

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Cancer Research

Tracking Bispecific Antibody-Induced T Cell Trafficking Using Luciferase-Transduced Human T Cells
Madelyn Espinosa-Cotton 1, Hong-Fen Guo 1, Nai-Kong V. Cheung 1
1Department of Pediatrics, Memorial Sloan Kettering Cancer Center

Here, we describe a method for transducing human T cells with luciferase to facilitate in vivo tracking of bispecific antibody-induced T cell trafficking to tumors in studies to evaluate the anti-tumor efficacy and mechanism of T cell-engaging bispecific antibodies.

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Cancer Research

Surgical Technique for Superior Cervical Ganglionectomy in a Murine Model
Qi Wang 1, Chun-Hao Chen 1, Hongbo Xu 2, Sylvie Deborde 1,3, Richard J. Wong 1,3
1Department of Surgery, Memorial Sloan Kettering Cancer Center, 2Department of Otolaryngology-Head and Neck Surgery, Beijing Tongren Hospital, Capital Medical University, 3David M. Rubenstein Center for Pancreatic Cancer Research, Memorial Sloan Kettering Cancer Center

The present protocol describes a mouse model of the ablation of adrenergic innervation by identifying and resecting the superior cervical ganglion.

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