This fourth edition explores fully up-to-date standardly used cryopreservation, vitrification, and freeze-drying protocols for specimens that are used for research purposes, conservation of genetic reserves, and applications in agriculture and medicine. Beginning with a section on the fundamentals as well as the use of mathematical modeling to solve cryobiological problems, the book continues with sections on technological aspects of freezing and drying, analytical methods to study protectant loading of cells and tissues, cell behavior during freezing and drying, and thermodynamic properties of preservation solutions, as well as cryopreservation, vitrification, and freeze-drying protocols for a wide variety of samples and different applications. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.

Part I: Fundamental Aspects of Cryopreservation and Freeze-Drying

1. Principles Underlying Cryopreservation and Freeze-Drying of Cells and Tissues

Willem F. Wolkers and Harriëtte Oldenhof

2. Principles of Ice-Free Cryopreservation by Vitrification

Gregory M. Fahy and Brian Wowk

3. The Principles of Freeze-Drying and Application of Analytical Technologies

Kevin R. Ward and Paul Matejtschuk

4. Mathematical Modeling and Optimization of Cryopreservation in Single Cells

James D. Benson

5. Mathematical Modeling of Protectant Transport in Tissues

Ross M. Warner and Adam Z. Higgins

Part II: Technologies and Methods to Study Freezing and Drying

6. Freezing Technology: Control of Freezing, Thawing, and Ice Nucleation

Peter Kilbride and Julie Meneghel

7. Microwave- and Laser-Assisted Drying for the Anhydrous Preservation of Biologics

Shangping Wang, Susan Trammell, and Gloria D. Elliott

8. High-Speed Video Cryomicroscopy for Measurement of Intracellular Ice Formation Kinetics

Jens O.M. Karlsson

9. Use of Ice Recrystallization Inhibition Assays to Screen for Compounds that Inhibit Ice Recrystallization

Anna A. Ampaw, August Sibthorpe, and Robert N. Ben

10. DSC Analysis of Thermophysical Properties for Biomaterials and Formulations

Wendell Q. Sun

11. Osmometric Measurements of Cryoprotective Agent Permeation into Tissues

Kezhou Wu, Leila Laouar, Nadia Shardt, Janet A.W. Elliott, and Nadr M. Jomha

12. Use of X-Ray Computed Tomography for Monitoring Tissue Permeation Processes

Ariadna Corral, Alberto Olmo, and Ramón Risco

13. Use of In Situ Fourier Transform Infrared Spectroscopy in Cryobiological Research

Willem F. Wolkers and Harriëtte Oldenhof

14. Raman Cryomicroscopic Imaging and Sample Holder for Spectroscopic Subzero Temperature Measurements

Guanglin Yu, Rui Li, and Allison Hubel

Part III: Cryopreservation and Freeze-Drying Protocols

15. Cryopreservation of Semen from Domestic Livestock: Bovine, Equine, and Porcine Sperm

Harriëtte Oldenhof, Willem F. Wolkers, and Harald Sieme

16. Cryopreservation of Avian Semen

17. Cryopreservation of Mouse Sperm for Genome Banking

Yuksel Agca and Cansu Agca

18. Cryopreservation of Marine Invertebrates: From Sperm to Complex Larval Stages

Estefania Paredes, Pablo Heres, Catarina Anjos, and Elsa Cabrita

19. Aseptic Cryoprotectant-Free Vitrification of Human Spermatozoa by Direct Dropping into a Cooling Agent

Mengying Wang, Evgenia Isachenko, Gohar Rahimi, Peter Mallmann, and Vladimir Isachenko

20. Cryopreservation of Mammalian Oocytes: Slow Cooling and Vitrification as Successful Methods for Cryogenic Storage

Victoria Keros and Barry J. Fuller

21. Vitrification of Porcine Oocytes and Zygotes in Microdrops on a Solid Metal Surface or Liquid Nitrogen

Tamas Somfai and Kazuhiro Kikuchi

22. Cryopreservation and Transplantation of Laboratory Rodent Ovarian Tissue for Genome Banking and Biomedical Research

Yuksel Agca and Cansu Agca

Jana Liebenthron and Markus Montag

24. Vitrification: A Simple and Successful Method for Cryostorage of Human Blastocysts

Juergen Liebermann

25. Vitrification of Equine In Vivo-Derived Embryos, after Blastocoel Aspiration

Carolina Herrera

26. Frozen Blood Reserves

Johan W. Lagerberg

27. Isolation, Cryopreservation, and Characterization of iPSC-Derived Megakaryocytes

Denys Pogozhykh, Rainer Blasczyk, and Constança Figueiredo

28. Chemically Defined, Clinical-Grade Cryopreservation of Human Adipose Stem Cells

Melany López and Ali Eroglu

29. Chemically-Defined and Xeno-Free Cryopreservation of Human Induced Pluripotent Stem Cells

Juliette Seremak and Ali Eroglu

30. Protocol for Cryopreservation of Endothelial Monolayers

Leah A. Marquez-Curtis, Nasim Eskandari, Locksley E. McGann, and Janet A.W. Elliott

31. Vitrification of Heart Valve Tissues

Kelvin G.M. Brockbank, Zhenzhen Chen, Elizabeth D. Greene, and Lia H. Campbell

32. Cryopreservation of Algae

Estefania Paredes, Angela Ward, Ian Probert, Léna Gouhier, and Christine N. Campbell

33. Cryopreservation of Fern Spores and Pollen

Anna Nebot, Victoria J. Philpott, Anna Pajdo, and Daniel Ballesteros

34. Cryopreservation of Plant Cell Lines Using Alginate Encapsulation

Heinz Martin Schumacher, Martina Westphal, and Elke Heine-Dobbernack

35. Cryopreservation of Plant Shoot Tips of Potato, Mint, Garlic, and Shallot Using Plant Vitrification Solution 3

Angelika Senula and Manuela Nagel

36. Cryopreservation of Seeds and Seed Embryos in Orthodox, Intermediate, and Recalcitrant Seeded Species

Daniel Ballesteros, Natalia Fanega-Sleziak, and Rachael Davies

37. Freeze-Drying of Proteins

Baolin Liu and Xinli Zhou

38. Freeze-Drying of Lactic Acid Bacteria: A Stepwise Approach for Developing a Freeze-Drying Protocol Based on Physical Properties

Fernanda Fonseca, Amélie Girardeau, and Stéphanie Passot

39. Preservation of Mammalian Sperm by Freeze-Drying

Levent Keskintepe and Ali Eroglu

40. Freeze-Drying of Decellularized Heart Valves for Off-the-Shelf Availability

Willem F. Wolkers and Andres Hilfiker