Animal Cell Electroporation and Electrofusion Protocols

ISBN-13: 9781489940612 / Angielski / Miękka / 2013 / 370 str.

Jac Nickoloff

Opis

The ability to introduce macromolecules into animal cells, includ- ing DNA, RNA, proteins, and other bioactive compounds has facili- tated a broad range of biological studies, from biochemistry and biophysics to molecular biology, cell biology, and whole animal stud- ies. Gene transfer technology in particular will continue to play an essential role in studies aimed at improving our understanding of the relationships between the gene structure and function, and it has impor- tant practical applications in both biotechnology and biomedicine, as evidenced by the current intense interest in gene therapy. Although DNA and other macromolecules may be introduced into cells by a variety of methods, including chemical treatments and microinjection, el- troporation has proven to be simpler to perform, more efficient, and effective with a wider variety of cell types than other techniques. The early and broad success of electric field-mediated DNA transfer soon prompted researchers to investigate electroporation for transferring other types of molecules into cells, including RNA, enzymes, antibodies, and analytic dyes. Animal Cell Electroporation and Electrofusion Protocols begins with three chapters that describe the theoretical and practical aspects of electroporation, including a review of the commercially available instrumentation. These introductory chapters will be of particular inter- est to those new to electric field technologies and to those developing protocols for as yet untested species or cell types. Nineteen chapters follow that present well-tested protocols for electroporation of proteins and DNA into insect, fish, and mammalian cells.

Recenzje

"This volume should fulfill its aim, to "take the guesswork out of experimental trials." -Society for General Microbiology Quarterly

"The book is very up-to-date, for instance it cover the most important recent developments in experiments with isolation of tumor suppressor genes in melanoma by cDNA microarray. The book represents an outstanding work, which should not be missed in all laboratories dealing with melanoma as well as in all departments where melanoma patients are diagnosed and treated. Those people who are interested to learn how the completion of the Human Genome Project significantly has been contributing to new progress in cancer management, exampled by human malignant melanoma, will certainly enjoy reading this publication as well." - Neoplasma

Spis treści

Part I. Theory and Instrumentation. Electroporation Theory: Concepts and Mechanisms. Effects of Pulse Length and Strength on Electroporation Efficiency. Instrumentation. Part II. Electroporation Protocols. The Introduction of Proteins into Mammalian Cells by Electroporation. Electroporation of Antigen-Presenting Cells for T-Cell Recognition and Cytotoxic T-Lymphocyte Priming. Electroporation of Antibodies into Mammalian Cells. Electroporation of Adherent Cells In Situ for the Introduction of Nonpermeant Molecules. Electrotransformation of Chinese Hamster Ovary Cells. Electroporation of Rat Pituitary Cells. Electroporation of Plasmid DNA into Normal Human Fibroblasts. Electroporation-Mediated Gene Transfer into Hepatocytes. Electroporation of Human Lymphoblastoid Cells. The Use of Electroporated Bovine Spermatozoa to Transfer Foreign DNA into Oocytes. Electroporation of Embryonic Stem Cells for Generating Transgenic Mice and Studying In Vitro Differentiation. Electrotransfection with 'Intracellular' Buffer. Effect of Cis-Located Human Satellite DNA on Electroporation Efficiency. Quantitation of Transient Gene Expression. Stable Integration of Vectors at High Copy Number for High-Level Expression in Animal Cells. Electroporation of Drosophila Embryos. Transformation of Fish Cells and Embryos. Electroporation of Cardiac Cells. Electroporation for Gene Therapy. Part III. Electrofusion Protocols. Electrofusion of Mammalian Cells. Stabilizing Antibody Secretion of Human Epstein Barr Virus-Activated B-Lymphocytes with Hybridoma Formation by Electrofusion. Electrofusion of Mammalian Oocytes and Embryonic Cells. Nuclear Transfer in Bovine Embryos. Electrofusion of Mouse Embryos to Produce Tetraploids. Spectrofluorometric Assay for Cell-Tissue Electrofusion. Cytometric Detection and Quantitation of Cell-Cell Electrofusion Products. Index.