Faecal Microbiota Sample Preparation with Cryoprotectants
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Solution Overview
Problem
Current faecal microbiota transplantation methods are empirical, lack preservation of anaerobic bacteria viability, and have variable effectiveness, requiring multiple treatments and pathogenic germ testing, especially for industrial-scale implementation and treating intestinal dysbioses like Clostridium difficile infections.
Innovation Solution
A method involving rapid sampling, storage in an oxygen-tight device under anaerobiosis, mixing with a saline solution containing cryoprotectants and bulking agents, optional filtration, and freezing at specific temperatures to preserve bacterial viability, facilitating safe and effective faecal microbiota transplantation.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional empirical transplantation methods are used, then the process is simple to implement, but the viability of anaerobic bacteria is not preserved and effectiveness is variable
Solution Approach 1:
The patent applies preliminary action by preparing the faecal sample with cryoprotectants and bulking agents before transplantation, and by pre-freezing the sample in oxygen-tight containers. This preliminary preparation ensures bacterial viability is preserved during storage and transport, eliminating the need for complex live-bacteria maintenance during transplantation while improving reliability
Solution Approach 2:
The patent uses oxygen-tight containers to create an inert anaerobic environment for sample collection, storage, and transport. This prevents oxidation and death of anaerobic bacteria, significantly improving their viability without requiring complex active preservation systems during the transplantation process
2Reliability
If multiple treatments are administered to ensure effectiveness, then the treatment success rate improves, but the treatment time and number of procedures increase
Solution Approach 1:
The patent prepares the sample in advance with optimal cryoprotectant concentrations and bulking agents, then freezes it for long-term storage. This preliminary optimization ensures that when transplantation occurs, the bacteria are already in maximum viability state, increasing treatment effectiveness in a single session and reducing the need for multiple repeat treatments
3Duration of action of stationary object
If faecal samples are stored without cryoprotectants, then the storage process is simple, but the bacterial viability deteriorates during storage
Solution Approach 1:
The patent changes the chemical parameters of the storage medium by adding cryoprotectants (such as glycerol, DMSO, or sugars) and bulking agents to the faecal sample before freezing. This chemical modification protects bacterial cell structures during freezing and storage, maintaining viability over extended periods without requiring ultra-low temperature storage infrastructure
Solution Approach 2:
The patent introduces cryoprotectants as intermediary substances between the bacteria and the freezing process. These intermediaries prevent ice crystal formation and cellular damage during freezing, enabling long-term storage while preserving bacterial viability without requiring complex cryogenic equipment
4Object-affected harmful factors
If pathogenic germ testing is performed on donor faeces, then the safety of transplantation improves, but the testing time and complexity increase
Solution Approach 1:
The patent extracts and concentrates potential pathogenic germs from the donor faecal sample through filtration and centrifugation steps before transplantation. This concentration step allows for more efficient and targeted pathogen detection using standard testing methods, improving safety assessment without requiring overly complex testing infrastructure
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The method ensures the viability of anaerobic bacteria, enhances transplantation efficacy, and provides a safe, easy-to-implement process for industrial-scale use, effectively treating intestinal dysbioses and infections like Clostridium difficile by maintaining microbial diversity and stability.
Implementation Method 1
placing said sample obtained in a) in an oxygen-tight collecting device, steps b) to e) being carried out under anaerobiosis
Implementation Method 2
mixing the sample obtained in b) with at least one saline aqueous solution comprising at least one cryoprotectant
Implementation Method 3
storing the mixture obtained in c) or d) by freezing at a temperature between −15° C. and −100° C., preferably between −60° C. and −90° C.
Data Source
AI summary
The present invention relates to a method for preparing a fecal 5 micro biota sample of a donor subject. Said method includes the following steps: a) collecting at least one fecal microbiota sample from the donor subject; b) within a period of less than 5 minutes after collecting the sample, placing said sample obtained in step a) in an oxygen-tight collection device; c) mixing the sample obtained in step b) with at least one aqueous saline IO solution containing at least one cryoprotectant and/or a filling agent; d) optionally, filtering the mixture obtained in step c), in particular by means of a filter comprising pores having a diameter of less than or equal to 0.7 mm, preferably less than or equal to 0.5 mm; and e) storing the mixture obtained in step c) or d) by freezing said mixture at a temperature 15 of between −15° C. and −100° C. Steps b) to e) are carried out in anaerobiosis.