The process of metastasis formation is hugely complex, as described in the introductory chapter of this book, and this complexity has led us to compile two volumes of methods, from a vastly divergent background that attempts to encompass the whole spectrum of cancer biology. This first volume, Metastasis Research Protocols: Analysis of Cells and Tissues, concentrates on analysis and mapping of molecules produced by cells and tissues and analysis of the molecular biology underlying their expression, whereas the second volume, Metastasis Research Protocols: Cell Behavior In Vitro and In Vivo,focuses sharply on the determination of cell behavior in vitro and in vivo. We have deliberately included chapters describing well-established and familiar te- niques (for example, SDS-PAGE and Western blotting [Chapter 11], and immunocytochemistry [Chapter 2]) in addition to the newer and more speci- ized approaches and specific examples of their application, because-although the methodology is readily available in the published literature and established in many laboratories-we wished these volumes to "stand alone" and to make accessible here the standard techniques that underpin much metastasis research for both the newcomer to the field and the seasoned researcher. Undoubtedly, owing to the complexity of the metastatic cascade and the wealth of research techniques involved in scientific approaches to its unraveling, and despite our best efforts to make these volumes as comprehensive as seems feasible, this is a tall order, and there will inevitably be omissions. For these we apologize.
Part I. Detection Methods for Cellular Markers of Metastatic Potential Histopathological Assessment of Metastasis Derek E. Roskell and Ian D. Buley Basic Immunocytochemistry for Light Microscopy Susan A. Brooks Multiple Labeling Techniques for Fluorescence Microscopy Amanda J. Atherton and Catherine Clarke Lectin Histochemistry to Detect Altered Glycosylation in Cells and Tissues Susan A. Brooks and Debbie M. S. Hall Immunocytochemical Detection and Characterization of Individual Micrometastatic Tumor Cells Stephan Braun and Klaus Pantel Understanding Metastasis Through Integrin Expression Lawrence John and Massimo Pignatelli Surrogate Markers of Angiogenesis and Metastasis Girolamo Ranieri and Giampietro Gasparini Measurement of Microvessel Density in Primary Tumors Helen E. Turner and Adrian L. Harris Assessment of Cellular Proliferation by Calculation of Mitotic Index and by Immunohistochemistry Cheryl E. Gillett Flow Cytometric Measurement of Cell Proliferation Richard S. Camplejohn SDS-PAGE and Western Blotting Techniques Christine Blancher and Adam Jones Assessment of Gelatinases (MMP-2 and MMP-9) by Gelatin Zymography Marta Toth and Rafael Fridman Part II. Genetic Aspects of Metastasis In Situ Hybridization to Localize mRNAs Richard Poulsom Fluorescence In Situ Hybridization Hakan Goker and Janet Shipley Comparative Genomic Hybridization Rebecca Roylance RNA Purification and Quantification Methods Adam Jones, Chisato Fujiyama, and Kenneth Smith Northern and Southern Blotting and the Polymerase Chain Reaction to Detect Gene Expression Thelma Tennant, Marina Chekmareva, and Carrie W. Rinker-Schaeffer Methylation Analysis of CpG Islands Richard Lilischkis, Hermann Kneitz, and Hans Kreipe Detection of Metastatic Tumor Cells in Blood by Reverse Transcriptase-Polymerase Chain Reaction Katja Haack, Krishna Soondrum, and Michael Neumaier Differential Display Ruey Ho Kao and Giulio Francia Part III. Mathematical Modeling of Metastases Predictive Mathematical Modeling in Metastasis Jonathan A. Sherratt