Hematopoietic Stem Cell Protocols

1. Front Matter

   Pages i-xi

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2. Isolation and Analysis of Hematopoietic Stem Cells from Mouse Embryos

   • Elaine Dzierzak, Marella de Bruijn

   Pages 1-14

3. The Purification of Mouse Hematopoietic Stem Cells at Sequential Stages of Maturation

   • Sean J. Morrison

   Pages 15-28

4. Flow Cytometry and Immunoselection of Human Stem Cells

   • Terry E. Thomas, Sara J. Abraham, Peter M. Lansdorp

   Pages 29-57

5. Method for Purification of Human Hematopoietic Stem Cells by Flow Cytometry

   • Michael J. Reitsma, Brenda R. Lee, Nobuko Uchida

   Pages 59-77

6. Noninvasive Measurement of Hematopoietic Stem Cell Cycling and Turnover by In Vivo Bromodeoxyuridine Incorporation

   • Gillian B. Bradford, Ivan Bertoncello

   Pages 79-92

7. Isolation and Characterization of Primitive Hematopoietic Cells Based on Their Position in the Cell Cycle

   • Edward F. Srour, Craig T. Jordan

   Pages 93-111

8. Hematopoietic Colony-Forming Cells

   • Makio Ogawa, Anne G. Livingston

   Pages 113-122

9. Long-Term Culture-Initiating Cell Assays for Human and Murine Cells

   • Cindy L. Miller, Connie J. Eaves

   Pages 123-141

10. The Cobblestone-Area-Forming Cell Assay

    • Gerald de Haan, Rob Ploemacher

    Pages 143-151

11. CFU-S

    • Ernest A. McCulloch

    Pages 153-160

12. Intrathymic Injection for Analysis of T-Cell Progenitor Activity

    • Libuse Jerabek, Irving L. Weissman

    Pages 161-165

13. Quantitation of Murine and Human Hematopoietic Stem Cells by Limiting-Dilution Analysis in Competitively Repopulated Hosts

    • Stephen J. Szilvassy, Franck E. Nicolini, Connie J. Eaves, Cindy L. Miller

    Pages 167-187

14. Ex Vivo Expansion of Human and Murine Hematopoietic Stem Cells

    • Cindy L. Miller, Julie Audet, Connie J. Eaves

    Pages 189-208

15. Hematopoietic Development of ES Cells in Culture

    • Gordon M. Keller, Saiphone Webb, Marion Kennedy

    Pages 209-230

16. Genetic Modification of Murine Hematopoietic Stem Cells by Retroviruses

    • Christian P. Kalberer, Jennifer Antonchuk, R. Keith Humphries

    Pages 231-242

17. Retroviral Transduction of FACS-Purified Hematopoietic Stem Cells

    • Claudiu V. Cotta, C. Weissman, Irving L. Weissman, Christopher A. Klug

    Pages 243-252

18. Retroviral-Mediated Transduction and Clonal Integration Analysis of Human Hematopoietic Stem and Progenitor Cells

    • Mo A. Dao, Jan A. Nolta

    Pages 253-274

19. Production of Lentiviral Vector Supernatants and Transduction of Cellular Targets

    • Richard E. Sutton

    Pages 275-286

20. Reverse Transcriptase-PCR Analysis of Gene Expression in Hematopoietic Stem Cells

    • Donald Orlic

    Pages 287-299

The ability to highly purify and characterize hematopoietic stem cells (HSC) from mice and humans has opened up an exceedingly rich field of basic science research with enormous clinical potential. Many of the techniques used in st- ies of HSC biology have become more standardized over the last several years, which makes it possible to compile a set of methods that can be used by both seasoned investigators and novices in the stem cell field. We have attempted to be as comprehensive as possible and yet focus on what we perceive to be the most widely used approaches for studies of murine and human HSC. This first edition of Hematopoietic Stem Cell Protocols will therefore have some obvious omissions that were dictated by contemporary circumstances. It is our hope that readers will feel free to contribute their personal suggestions for further chapters as well as on how existing chapters can be improved for future editions. We certainly expect that old approaches will be refined, new assays will be developed, and other animal model and vector systems will be described that will become the new gold standards for future work. Our s- cere thanks goes out to all of the contributors and to those in the stem cell field that have enlarged our thinking and provided new tools to further understand this fascinating cell type.

"Knowledge of hematopoietic stem cells is crucial in the development of clinical medicine, especially hematopoietic stem cell transplantation, solid organ transplantation, tissue engineering for the treatment of damaged tissue. Research in the area has made major advance in the past two decades. Various technologies were produced and contributed to the development. This book describes detailed techniques in each experiment as if authors are teaching to the investigators by their side. Each section also contains notes and introductions that explain practical tips, historical and logical aspects of those experiments. This book covers current important methodologies in hematopoietic stem cell research and helps investigators to perform correct techniques excellently. "- Doody's Health Sciences Book Review Journal. Numerical Score: 91 - 4 Stars!

"The book presents intimate knowledge of stem cell technology and useful practical guide through laboratory protocols. Despite detailed step by step protocols with plenty of valuable pieces of information about critical assay points, book offers even a comprehensive biological and molecular background of HSC. Essential for novices and inspiring for experienced investigators, Hematopoietic Stem Cell Protocols, will not have a fate of book forgotten at the upper bookcase shelf but can accomplish the idea of information source easily reached at your bench." The book is well referenced, and readers will easily be able to locate primary literature for in-depth reading on specific topics. The figures and illustrations are extremely useful in summarizing complex topics that are discussed." -Neoplasma

"Comprehensive and state-of-the-art, Hematopoietic Stem Cell Protocols offers both novice and experienced investigators alike an invaluable compendium of highly practical techniques for studying a wide variety of stem cell topics, including blood formation, blood-based disease, and such blood cancers as leukemia." -Clinical Laboratory

• University of Alabama-Birmingham, Birmingham, AL

  Christopher A. Klug

• University of Kentucky, Lexington, KY

  Craig T. Jordan

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