Gnotobiotic Mouse Technology An Illustrated Guide

A Brief History of Germ-Free Life
References

Overview of Gnotobiotic Technology
Overview
How Do We Get Germ-Free Mice?
Germ-Free Housing and Barrier Types
References

Equipment and Terminology

Personal Protective Equipment
Introduction
Types of PPE
Applying Sterile PPE

Sterilants and Sterilization
Introduction
Sterilization Methods
Cold Sterilization
Wrapping Materials for Autoclaving
Gas Sterilization (Ethylene Oxide)

Isolator Setup
Introduction
Part 1: Small (5.5 x 3 feet) Isolator Assembly
Part 2: Large Isolator Assembly

Port Entry and Exit
Introduction
Supplies
Preparing, Opening, and Closing the Port
Proper Placement of Materials

Sterilizing Food and Supplies
Introduction
Biological Indicators
Assembling Supply Cylinders: Installing HEPA Filtration Media
Supplies
Prepare the Cylinder
Fill the Cylinder
Water Sterilization

Supply Cylinder Entry
Introduction
Supplies
Procedure

Isolator Maintenance
Introduction
Part 1: Cleaning and Maintenance
Part 2: Replacing Gloves on an Active Isolator
Part 3: Repairing Holes in Isolators
Part 4: Working in the Isolator

Aseptic Mouse Transfer
Introduction
Part 1: Internal Transfer between Isolators
Part 2: External Transfer of Mice

Working with Germ-Free or Gnotobiotic Mice in the Class II Biosafety Cabinet
Introduction
Supplies
Experimental Manipulations in the Biosafety Cabinet

Shipping Mice
Introduction
Procedure

Rederivation
Introduction
Derivation by Cesarean Section
Derivation by Embryo Transfer
References

Microbiological Testing
Introduction
Serologic Screening
Part 1: Bacterial Culture
Part 2: Examination of Gram-Stained Slides for the Presence of Bacteria
References

Genetic Testing
References

Record Keeping
Introduction
Colony Management Documents
Project-Planning Documents
Other Documents

Facility Setup and Management
Introduction
Establishing a New Facility

Appendix: Sources for Equipment and Supplies

Biography

Chriss J. Vowles co-manages a multi-investigator germ-free research laboratory at the University of Michigan in Ann Arbor. He began his career at the University of Michigan in 2003, working full-time as a husbandry technician in the Unit for Laboratory Animal Medicine. In 2006, he discovered gnotobiotic technology. At that time, the Germ Free Laboratory was just starting. Chriss joined the research group on the ground floor and has been growing with it ever since. The idea of maintaining a germ-free, complex organism fascinated him then and still captivates him today. After eight years of maintaining a germ-free colony of mice, the laboratory is still evolving, he is still learning, and his trials, tribulations, and rewards are constant. Natalie E. Anderson is a research technician lead at the University of Michigan's Germ Free Core. In 2009, she graduated from Michigan State University with a bachelor's degree in animal science and accepted a position with SoBran's contract for the National Institute of Allergy and Infectious Diseases (NIAID). In December 2010, she returned to Michigan to take a position with the University of Michigan's Unit for Lab Animal Medicine. Shortly thereafter, she started working part-time in the Germ Free Core and quickly learned to love the daily challenges involved in maintaining a germ-free colony. Natalie joined the core full time in July 2013. Kathryn A. Eaton has been working with germ-free and gnotobiotic animal models for 30 years. She started her gnotobiology career at Ohio State University, where she did her PhD research on Helicobacter pylori in gnotobiotic piglets. She went on to study mice several years later, and in 2004 she established the University of Michigan Germ Free Mouse Laboratory, which she now directs. Dr. Eaton is a board-certified veterinary pathologist with research interests in bacterial enteric disease and immunology. In addition to H. pylori ,

"This book introduces and describes techniques utilized for the production, maintenance, and experimental manipulation of germ-free and gnotobiotic mice in a research support facility. The introductory chapter provides history and identifies animals that were successfully derived and maintained in a germ-free setting. …

The middle portion of the book provides detailed information regarding isolator setup. … provides step-by-step descriptions and illustrations for assembly, inflation, sterilization, and decontamination….

The next portion of the book describes the proper technique for port entry and exit, isolator maintenance, and the use of supply cylinders and supply isolators.

The final chapters of the book discuss methods of microbiological testing such as the Gram stain procedure and quantitative PCR. … Genetic testing and record keeping are discussed. … Chapter 18 describes facility setup and management of maintaining a gnotobiotic laboratory. Gnotobiotic laboratories are labor intensive and extremely expensive. Funding options and space requirements when establishing and/or maintaining an existing gnotobiotic laboratory must be discussed with institutional directors.

Overall, this book is reasonably priced and available in electronic and paperback versions. The combination of procedural descriptions and detailed illustrations are an advantage of this book. The chapter describing isolator setup is very descriptive with complementary illustrations. This book serves as a good reference for new and/or existing gnotobiotic facilities."

Dondrae Coble, DVM, DACLAM in Laboratory Animal Practitione