Related Experiment Video
Updated: Jan 23, 2026
Murine Fecal Isolation and Microbiota Transplantation
Published on: May 26, 2023
A Guide for Ex Vivo Handling and Storage of Stool Samples Intended for Fecal Microbiota Transplantation
Sebastian D Burz1,2, Anne-Laure Abraham3, Fernanda Fonseca4
1Micalis Institute, INRA, AgroParisTech, Université Paris-Saclay, 78350, Jouy-en-Josas, France.
Abstract:
Owing to the growing recognition of the gut microbiota as a main partner of human health, we are expecting that the number of indications for fecal microbiota transplantation (FMT) will increase. Thus, there is an urgent need for standardization of the entire process of fecal transplant production. This study provides a complete standardized procedure to prepare and store live and ready-to-use transplants that meet the standard requirements of good practices to applied use in pharmaceutical industry. We show that, if time before transformation to transplants would exceed 24 hours, fresh samples should not be exposed to temperatures above 20 °C, and refrigeration at 4 °C can be a safe solution. Oxygen-free atmosphere was not necessary and simply removing air above collected samples was sufficient to preserve viability. Transplants prepared in maltodextrin-trehalose solutions, stored in a -80 °C standard freezer and then rapidly thawed at 37 °C, retained the best revivification potential as proven by 16S rRNA profiles, metabolomic fingerprints, and flow cytometry assays over a 3-month observation period. Maltodextrin-trehalose containing cryoprotectants were also efficient in preserving viability of lyophilized transplants, either in their crude or purified form, an option that can be attractive for fecal transplant biobanking and oral formulation.
Insights
Standardized fecal microbiota transplantation (FMT) production ensures viability. Optimal storage involves maltodextrin-trehalose solutions at -80°C, preserving microbial function for 3 months.
Area of Science:
- Microbiology
- Gastroenterology
- Biotechnology
Background:
- The gut microbiota plays a crucial role in human health.
- Increasing indications for fecal microbiota transplantation (FMT) necessitate standardized production protocols.
- Current FMT production lacks universally accepted standardization, impacting clinical application.
Purpose of the Study:
- To establish a standardized procedure for preparing and storing fecal microbiota transplantation (FMT) products.
- To ensure the viability and quality of FMT products for pharmaceutical industry standards.
- To evaluate different storage conditions and cryoprotective agents for FMT preservation.
Main Methods:
- Development of a standardized protocol for FMT preparation and storage.
- Assessment of sample viability under various temperature conditions (e.g., 4°C, 20°C) and atmospheric controls.
- Evaluation of cryoprotective agents (maltodextrin-trehalose) for preserving FMT viability during freezing (-80°C) and lyophilization.
- Analysis of microbial viability using 16S rRNA profiling, metabolomic fingerprinting, and flow cytometry over a 3-month period.
Main Results:
- Refrigeration at 4°C is suitable for samples stored <24 hours; temperatures >20°C should be avoided.
- An oxygen-free atmosphere is not required; simple air removal suffices for viability.
- FMT prepared with maltodextrin-trehalose, stored at -80°C, and rapidly thawed at 37°C showed optimal viability over 3 months.
- Maltodextrin-trehalose effectively preserved viability in both crude and purified lyophilized FMT.
Conclusions:
- A standardized FMT production and storage protocol has been developed, meeting good practice requirements.
- Maltodextrin-trehalose cryoprotectants are effective for preserving FMT viability, supporting long-term biobanking and potential oral formulations.
- The findings facilitate the pharmaceutical application of FMT by ensuring product quality and stability.
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