A collection of both well-established and cutting-edge methods for investigating breast cancer biology not only in the laboratory, but also in clinical settings. These readily reproducible techniques solve a variety of problems, ranging from how to collect, store, and prepare human breast tumor samples for analysis, to analyzing cells in vivo and in vitro. Additional chapters address the technology of handling biopsies, new methods for analyzing genes and gene expression, markers of clinical outcome and progress, analysis of tumor-derived proteins and antigens, validating targets, and investigating the biology of newly discovered genes.
Part I. Preparation of Cell and Tissue Samples Breast Tissue Banking: Collection, Handling, Storage, and Release of Tissue for Breast Cancer Research Linda Snell and Peter H. Watson Frozen Biopsy Collection and Storage: Frozen Biopsy Samples Russell Leek Serial Biopsies/Fine-Needle Aspirates and Their Assessment Irene Boeddinghaus and Stephen R. D. Johnson Breast Tissue Microarrays Robert J. Springall and Cheryl E. Gillett Preparation of Tumor Homogenates for Subsequent Preparation of Cytosols, Membrane Fractions, RNA, and DNA Naomi Robertson and Russell Leek Isolation of RNA From Tumor Samples: Single-Step Guanidinium Acid-Phenol Method Naomi Robertson and Russell Leek Isolation of DNA From Tumor Samples Naomi Robertson and Russell Leek Laser-Assisted Microdissection and Isolation of DNA and RNA Ulrich Lehmann and Hans Kreipe Part II. Markers of Clinical Outcome and Prognosis Traditional and Established Indicators of Prognosis and Treatment Success Derek E. Roskell and Ian D. Buley Sentinel Lymph Node Biopsy in Breast Cancer Celia Chao and Kelly M. McMasters Pathological Evaluation of Axillary Sentinel Lymph Nodes in Breast Cancer Michael Z. Gilcrease and Aysegul Sahin Measurement of Estrogen Receptor Status by Immunocytochemistry in Paraffin Wax Sections Bharat Jasani, Anthony Douglas-Jones, Anthony Rhodes, Susan Wozniak, Peter J. Barrett-Lee, Julia Gee, and Robert Nicholson Markers of Apoptosis Fazlul H. Sarkar and Yiwei Li Quantitative Angiogenesis in Breast Cancer Stephen B. Fox Part III. Analysis of Tumor-Derived Proteins and Antigens Immunohistochemistry Cheryl E. Gillett Detection of Aberrant Glycosylation in Breast Cancer Using Lectin Histochemistry Tracey M. Carter and Susan A. Brooks SDS-PAGE and Western Blotting to Detect Proteins and Glycoproteins of Interest in Breast Cancer Research Chloe Osborne and Susan A. Brooks Breast Cancer Proteomics Using Two-Dimensional Electrophoresis: Studying the Breast Cancer Proteome Miriam V. Dwek and Sarah L. Rawlings Procedures for the Quantitative Protein Determination of Urokinase and Its Inhibitor, PAI-1, in Human Breast Cancer Tissue Extracts by ELISA Manfred Schmitt, Alexandra S. Sturmheit, Anita Welk, Christel Schnelldorfer, and Nadia Harbeck Part IV. Analysis of Genes and Gene Expression in Tumor Specimens Fluorescence In Situ Hybridization and Comparative Genomic Hybridization Patricia Gorman and Rebecca Roylance Fluorescence In Situ Hybridization Assessment of c-myc Gene Amplification in Breast Tumor Tissues Jan K. Blancato, Mary Steele Williams, and Robert B. Dickson Detection of HER2 Gene Amplification by Fluorescence In Situ Hybridization in Breast Cancer John M. S. Bartlett and Amanda Forsyth In Situ Hybridization Combined With Immunohistochemistry to Localize Gene Expression Rosemary Jeffery, Toby Hunt, and Richard Poulsom Quantitation of RNA by Ribonuclease Protection Assay John W. Moore Identification of Steroid Hormone-Regulated Genes in Breast Cancer Bruce R. Westley and Felicity E. B. May Sequencing of the Tumor Suppressor Gene TP 53 Barbro Linderholm, Torbjörn Norberg, and Jonas Bergh Expression Profiling Using cDNA Microarrays Chris Jones, Peter Simpson, Alan Mackay, and Sunil R. Lakhani Gene Expression Analysis Using Filter cDNA Microarrays Peter Simpson, Chris Jones, Alan Mackay, and Sunil R. Lakhani Part V. Studying Cancer Cell Behavior In Vitro and In Vivo Methods to Analyze the Effects of the Urokinase System on Cancer Cell Adhesion, Proliferation, Migration,
Many new technologies involving large-scale analysis of gene expression, novel treatments, and emergent combined therapy approaches have opened fresh possibilities for understanding and effectively managing breast cancer. In Breast Cancer Research Protocols, active researchers and clinicians describe in detail both well-established and cutting-edge methods for investigating breast cancer biology not only in the laboratory, but also in clinical settings. These readily reproducible techniques solve a variety of problems, ranging from how to collect, store, and prepare human breast tumor samples for analysis, to analyzing cells in vivo and in vitro. Additional chapters address the technology of handling biopsies, new methods for analyzing genes and gene expression, markers of clinical outcome and progress, analysis of tumor-derived proteins and antigens, validating targets, and investigating the biology of newly discovered genes. The protocols follow the successful Methods in Molecular Medicine™ series format, each offering step-by-step laboratory instructions, an introduction outlining the principles behind the technique, lists of the necessary equipment and reagents, and tips on troubleshooting and avoiding known pitfalls.
Comprehensive and highly practical, Breast Cancer Research Protocols offers laboratory and clinical researchers the tools needed to illuminate breast cancer biology and advance the efficacy of its treatment today.
Preparation of Cell and Tissue Samples.- Breast Tissue Banking.- Frozen Biopsy Collection and Storage.- Serial Biopsies/Fine-Needle Aspirates and Their Assessment.- Breast Tissue Microarrays.- Preparation of Tumor Homogenates for Subsequent Preparation of Cytosols, Membrane Fractions, RNA, and DNA.- Isolation of RNA From Tumor Samples.- Isolation of DNA From Tumor Samples.- Laser-Assisted Microdissection and Isolation of DNA and RNA.- Markers of Clinical Outcome and Prognosis.- Traditional and Established Indicators of Prognosis and Treatment Success.- Sentinel Lymph Node Biopsy in Breast Cancer.- Pathological Evaluation of Axillary Sentinel Lymph Nodes in Breast Cancer.- Measurement of Estrogen Receptor Status by Immunocytochemistry in Paraffin Wax Sections.- Markers of Apoptosis.- Quantitative Angiogenesis in Breast Cancer.- Analysis of Tumor-Derived Proteins and Antigens.- Immunohistochemistry.- Detection of Aberrant Glycosylation in Breast Cancer Using Lectin Histochemistry.- SDS-PAGE and Western Blotting to Detect Proteins and Glycoproteins of Interest in Breast Cancer Research.- Breast Cancer Proteomics Using Two-Dimensional Electrophoresis.- Procedures for the Quantitative Protein Determination of Urokinase and Its Inhibitor, PAI-1, in Human Breast Cancer Tissue Extracts by ELISA.- Analysis of Genes and Gene Expression in Tumor Specimens.- Fluorescence In Situ Hybridization and Comparative Genomic Hybridization.- Fluorescence In Situ Hybridization Assessment of c-myc Gene Amplification in Breast Tumor Tissues.- Detection of HER2 Gene Amplification by Fluorescence In Situ Hybridization in Breast Cancer.- In Situ Hybridization Combined With Immunohistochemistry to Localize Gene Expression.- Quantitation of RNA by Ribonuclease Protection Assay.- Identification of Steroid Hormone-Regulated Genes in Breast Cancer.- Sequencing of the Tumor Suppressor Gene TP 53.- Expression Profiling Using cDNA Microarrays.- Gene Expression Analysis Using Filter cDNA Microarrays.- Studying Cancer Cell Behavior in Vitro and In Vivo.- Methods to Analyze the Effects of the Urokinase System on Cancer Cell Adhesion, Proliferation, Migration, and Signal Transduction Events.- Phospho-Specific Antibodies as a Tool to Study In Vivo Regulation of BRCA1 After DNA Damage.- Models of Hormone Resistance In Vitro and In Vivo.- Generation of Genetically Modified Embryonic Stem Cells for the Development of Knockout Mouse Animal Model Systems.- In Vivo Xenograft Models of Breast Cancer Metastasis.- Neoadjuvant Endocrine Therapy Models.- Primary Mouse Endothelial Cell Culture for Assays of Angiogenesis.
Inhaltsverzeichnis
Part I. Preparation of Cell and Tissue Samples
Breast Tissue Banking: Collection, Handling, Storage, and Release of Tissue for Breast Cancer Research
Linda Snell and Peter H. Watson
Frozen Biopsy Collection and Storage: Frozen Biopsy Samples
Russell Leek
Serial Biopsies/Fine-Needle Aspirates and Their Assessment
Irene Boeddinghaus and Stephen R. D. Johnson
Breast Tissue Microarrays
Robert J. Springall and Cheryl E. Gillett
Preparation of Tumor Homogenates for Subsequent Preparation of Cytosols, Membrane Fractions, RNA, and DNA
Naomi Robertson and Russell Leek
Isolation of RNA From Tumor Samples: Single-Step Guanidinium Acid-Phenol Method
Naomi Robertson and Russell Leek
Isolation of DNA From Tumor Samples
Naomi Robertson and Russell Leek
Laser-Assisted Microdissection and Isolation of DNA and RNA
Ulrich Lehmann and Hans Kreipe
Part II. Markers of Clinical Outcome and Prognosis
Traditional and Established Indicators of Prognosis and Treatment Success
Derek E. Roskell and Ian D. Buley
Sentinel Lymph Node Biopsy in Breast Cancer
Celia Chao and Kelly M. McMasters
Pathological Evaluation of Axillary Sentinel Lymph Nodes in Breast Cancer
Michael Z. Gilcrease and Aysegul Sahin
Measurement of Estrogen Receptor Status by Immunocytochemistry in Paraffin Wax Sections
Bharat Jasani, Anthony Douglas-Jones, Anthony Rhodes, Susan Wozniak, Peter J. Barrett-Lee, Julia Gee, and Robert Nicholson
Markers of Apoptosis
Fazlul H. Sarkar and Yiwei Li
Quantitative Angiogenesis in Breast Cancer
Stephen B. Fox
Part III. Analysis of Tumor-Derived Proteins and Antigens
Immunohistochemistry
Cheryl E. Gillett
Detection of Aberrant Glycosylation in Breast Cancer Using Lectin Histochemistry
Tracey M. Carter and Susan A. Brooks
SDS-PAGE and Western Blotting to Detect Proteins and Glycoproteins of Interest in Breast Cancer Research
Chloe Osborne and Susan A. Brooks
Breast Cancer Proteomics Using Two-Dimensional Electrophoresis: Studying the Breast Cancer Proteome
Miriam V. Dwek and Sarah L. Rawlings
Procedures for the Quantitative Protein Determination of Urokinase and Its Inhibitor, PAI-1, in Human Breast Cancer Tissue Extracts by ELISA
Manfred Schmitt, Alexandra S. Sturmheit, Anita Welk, Christel Schnelldorfer, and Nadia Harbeck
Part IV. Analysis of Genes and Gene Expression in Tumor Specimens
Fluorescence In Situ Hybridization and Comparative Genomic Hybridization
Patricia Gorman and Rebecca Roylance
Fluorescence In Situ Hybridization Assessment of c-myc Gene Amplification in Breast Tumor Tissues
Jan K. Blancato, Mary Steele Williams, and Robert B. Dickson
Detection of HER2 Gene Amplification by Fluorescence In Situ Hybridization in Breast Cancer
John M. S. Bartlett and Amanda Forsyth
In Situ Hybridization Combined With Immunohistochemistry to Localize Gene Expression
Rosemary Jeffery, Toby Hunt, and Richard Poulsom
Quantitation of RNA by Ribonuclease Protection Assay
John W. Moore
Identification of Steroid Hormone-Regulated Genes in Breast Cancer
Bruce R. Westley and Felicity E. B. May
Sequencing of the Tumor Suppressor Gene TP 53
Barbro Linderholm, Torbjörn Norberg, and Jonas Bergh
Expression Profiling Using cDNA Microarrays
Chris Jones, Peter Simpson, Alan Mackay, and Sunil R. Lakhani
Gene Expression Analysis Using Filter cDNA Microarrays
Peter Simpson, Chris Jones, Alan Mackay, and Sunil R. Lakhani
Part V. Studying Cancer Cell Behavior In Vitro and In Vivo
Methods to Analyze the Effects of the Urokinase System on Cancer Cell Adhesion, Proliferation, Migration,
Klappentext
A collection of both well-established and cutting-edge methods for investigating breast cancer biology not only in the laboratory, but also in clinical settings. These readily reproducible techniques solve a variety of problems, ranging from how to collect, store, and prepare human breast tumor samples for analysis, to analyzing cells in vivo and in vitro. Additional chapters address the technology of handling biopsies, new methods for analyzing genes and gene expression, markers of clinical outcome and progress, analysis of tumor-derived proteins and antigens, validating targets, and investigating the biology of newly discovered genes.
Includes supplementary material: sn.pub/extras
