Cold Spring Harbor Laboratory View Institution's Website 7 articles published in JoVE Cancer Research Establishment and Culture of Patient-Derived Breast Organoids Disha Aggarwal1,2, Suzanne Russo1, Payal Naik1, Sonam Bhatia1, David L. Spector1,2 1Cold Spring Harbor Laboratory, Cold Spring Harbor, 2Genetics Graduate Program, Stony Brook University A detailed protocol is provided here for establishing human breast organoids from patient-derived breast tumor resections or normal breast tissue. The protocol provides comprehensive step-by-step instructions for culturing, freezing, and thawing human patient-derived breast organoids. Cancer Research Longitudinal Intravital Imaging Through Clear Silicone Windows Laura Maiorino*1,2,3, Margaret Shevik*1,4,5, José M. Adrover1, Xiao Han1,6, Elias Georgas7,8, John Erby Wilkinson9, Harrison Seidner1, Leonie Foerschner1, David A. Tuveson1, Yi-Xian Qin8, Mikala Egeblad1 1Cold Spring Harbor Laboratory, 2Cold Spring Harbor Laboratory School of Biological Sciences, 3Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, 4Medical Scientist Training Program, School of Medicine, Stony Brook University, 5Graduate Program in Pharmacology, Stony Brook University, 6Graduate Program in Genetics, Stony Brook University, 7Graduate Program in Biomedical Engineering, Stony Brook University, 8Department of Biomedical Engineering, Stony Brook University, 9Department of Pathology, University of Michigan An approach is here presented for long-term intravital imaging using optically clear, silicone windows that can be glued directly to the tissue/organ of interest and the skin. These windows are cheaper and more versatile than others currently used in the field, and the surgical insertion causes limited inflammation and distress to the animals. Genetics A New Toolkit for Evaluating Gene Functions using Conditional Cas9 Stabilization Polona Šafarič Tepeš1,2, Raffaella Sordella1 1Cold Spring Harbor Laboratory, 2Faculty of Pharmacy, University of Ljubljana Here, we describe a genome-editing tool based on the temporal and conditional stabilization of clustered regularly interspaced short palindromic repeat- (CRISPR-) associated protein 9 (Cas9) under the small molecule, Shield-1. The method can be used for cultured cells and animal models. Neuroscience Single-cell RNA Sequencing of Fluorescently Labeled Mouse Neurons Using Manual Sorting and Double In Vitro Transcription with Absolute Counts Sequencing (DIVA-Seq) Anirban Paul1, Z. Josh Huang1 1Cold Spring Harbor Laboratory This protocol describes the manual sorting procedure to isolate single fluorescently labeled neurons followed by in vitro transcription-based mRNA amplification for high-depth single-cell RNA sequencing. Medicine Live Imaging of Drug Responses in the Tumor Microenvironment in Mouse Models of Breast Cancer Elizabeth S. Nakasone1,2, Hanne A. Askautrud2,3, Mikala Egeblad2 1Watson School of Biological Sciences, 2Cold Spring Harbor Laboratory, 3Departments of Medical Genetics, University of Oslo and Oslo University Hospital We describe a method for imaging response to anti-cancer treatment in vivo and at single cell resolution. Biology A Novel Bayesian Change-point Algorithm for Genome-wide Analysis of Diverse ChIPseq Data Types Haipeng Xing1, Willey Liao1,2, Yifan Mo1,2, Michael Q. Zhang2,3 1Department of Applied Mathematics & Statistics, Stony Brook University, 2Computational Biology and Bioinformatics, Cold Spring Harbor Laboratory, 3Department of Molecular and Cell Biology, University of Texas at Dallas Our Bayesian Change Point (BCP) algorithm builds on state-of-the-art advances in modeling change-points via Hidden Markov Models and applies them to chromatin immunoprecipitation sequencing (ChIPseq) data analysis. BCP performs well in both broad and punctate data types, but excels in accurately identifying robust, reproducible islands of diffuse histone enrichment. Biology In Situ Hybridization for the Precise Localization of Transcripts in Plants Marie Javelle1, Cristina F. Marco1, Marja Timmermans1 1Cold Spring Harbor Laboratory The in situ hybridization protocol described here allows a direct localization of mRNA and small RNA expression at the cellular level with high sensitivity and specificity. The procedure is optimized for paraffin-embedded plant tissue sections, is applicable to a wide range of plants and tissues, and can be completed within ten days.