Our Core Facility Resources

The Emory University Gnotobiotic Animal Facility is located in the newly constructed five-story, 200,000-square-foot Health Sciences Research Building (HSRB), situated in rooms next to the Specific Pathogen Free (MPF) murine housing, as well as the Transgenic Mouse and Gene Targeting Core. The facility contains a number of 3’ foot wide rigid isolators (Parkbio), each with the capacity to house 16 mice cages each. Class II biological cabinets are available to investigators for experimental use. In addition, the facility has a Tecniplast ISOcageP Bioexclusion system. These are airtight individual mouse cages with high positive pressure that are specifically designed for cage-scale germ-free, gnotobiotic and bioexclusion studies.

Animal welfare: The facility is run under Emory University’s IACUC approved Standard Operating Procedures which oversees and certifies that care and use of the animals is ethical and humane. The IACUC oversight is implemented by an ongoing review and approval of the facility by way of written animal use protocols submitted for review by IACUC committee members. Investigators wishing to use the facility will submit an amendment to their IACUC by attaching the SOP for the Gnotobiotic Animal Facility, and a description of the specific procedure they wish to undertake within the facility.

Murine strains bred and held within the facility: The facility currently has breeding colonies of germ-free C57BL/6J, and of with colonized with Altered Schaedler Flora (ASF) colonized C57BL/6J. Germ-free and ASF C57BL/6J mice are available for purchase from the facility.

Transferring new mouse strains into the facility: Currently, we accept re-derived germ-free mice from approved vendors. Mice must be shipped in approved transporters. Mice are transferred into newly set up isolators, and kept as the only genotype in said isolator for 4 weeks, and monitored weekly for germ-free status.

Transferring animals from the facility to laboratories within Emory University: Investigators will be responsible for transportation mice from the HSRB to other research buildings within Emory. Transport procedures are outlined in the IACUC website. Please contact the Gnotobiotic facility director or staff if you require further information regarding transport. 

Specialized diets: We have the capacity to rear mice on germ-free specialized diets including high fat diets or diets fortified with supplements. 

Microbiological Testing: Rigorous microbiological testing is conducted on all autoclaved material entering the isolators, including the food, water and bedding. We also regularly monitor germ-free mice within isolators and periodic control necropsies. Our tests include 1) real time quantitative PCR amplification of the V4 region 16S rRNA gene, 2) plating of fecal samples on BBL Brain Heart infusion with 10% Sheep Blood plates (nonselective controls for all strains) and incubation at 37ºC for 7 days, 3) Incubation in Thioglycollate medium with indicator, a medium used for the isolation and cultivation of aerobes, anaerobes and microaerophiles that are not excessively fastidious, and 4) Gram stain, which is a general visual examination for the presence of bacteria, and distinguishes between Gram-positive and Gram-negative organisms.

 

 

Gnotobiotic experiments in the Tecniplast Bioexclusion ISOcage system:  Precise modulation of the microbiome is increasingly appreciated as feasible approach to positively impact health and disease. However, heterogeneity of the microbiome between murine colonies housed in various facilities nationwide represent a tangible challenge to scientific rigor and reproducibility. Indeed, incongruent results have been observed between facilities when mice of the same phenotype were subjected to the same assays.

We have developed a facility to directly control for microbiome heterogeneity by, where feasible, undertaking our assays in mice gnotobiotically colonized them with a defined flora. We undertake this by housing our mice in within our cutting edge Tecniplast ISOcageP Bioexclusion system. These are airtight individual mouse cages with high positive pressure that are specifically designed for germ-free, gnotobiotic and bioexclusion studies. The ISO cage system is the latest design for gontobiotics and germ-free animals because it allows researchers to undertake up to 36 simultaneous gnotobiotic studies, compared to only one study at a time in conventional multi-cage gnotobiotic isolators.

For gnotobiotic experiments, mice can be initially raised under germ-free conditions. After weaning (3 weeks) mice may be transferred to an ISOcage cage. For experiments with a fully characterized controlled microbiome background, mice may be gnotobiotically colonized with Altered Schaedler Flora (ASF). An ASF-associated colony is maintained in the Emory Gnotobiotic Animal Facility. The colony was established by first raising germ-free C57BL/6 mice in an isolator. Thereafter, fecal pellets from mice harboring only ASF organisms were purchased from Taconic Biosciences and introduced into the isolators. Breeding pairs of mice that have been colonized with ASF from birth were set up and used to maintain the ASF colony. Monthly testing for the presence of the eight microbes in the ASF is undertaken by PCR using primers specific to each bacterial species. ASF is a standardized microbial blend of eight bacteria, which allows for the exquisitely meticulous studies in a fully defined microbiome background. Importantly, compared to germ-free animals, ASF-associated mice have normally developed immune system, they have resistance to opportunistic pathogens, and normal gastrointestinal health. They are thus an excellent representative model of normal mice with a defined microbiome.

Generating humanized mice within gnotobiotic cages:

As many as 85% of the sequences detected in the murine microbiome have not been detected in humans. Generating germ-free mice and then colonizing them with fecal material from human patients and donors overcomes this limitation. This approach maintains a microbiome diversity and profile similar to the human donor and thus represents a faithful model to examine colonization resistance in humans. “Humanized” germ-free mice are housed within our Tecniplast ISOcageP Bioexclusion system. For example, mice may be colonized with fecal material from diseased patients, and compared to mice colonized with the healthy control individuals.