High-Density Cryopreserved Cell Bank via Perfusion Culture
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Solution Overview
Problem
Conventional cell banking methods face challenges in achieving high cell densities for cryopreservation without damaging cells, as lower density stocks are inefficient for large volume cultures and centrifugation-based concentration methods are harmful to cells.
Innovation Solution
The method employs perfusion culture techniques coupled with non-centrifugal cell retention systems, specifically alternating tangential flow filtration, to concentrate cells to high densities while maintaining cell viability, followed by cryopreservation with DMSO addition.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If centrifugation-based concentration methods are used to increase cell density, then cell density is improved, but cell viability deteriorates due to mechanical damage
Solution Approach 1:
The patent replaces the mechanical centrifugation system with a perfusion culture system that uses controlled medium flow to achieve cell concentration. Instead of applying centrifugal force that damages cells, the system uses a bioreactor with a filter to separate and concentrate cells through fluid flow, eliminating mechanical cell damage while achieving high cell density for cryopreservation
Solution Approach 2:
The patent introduces a filter as an intermediary component between the cell culture medium and the concentration process. The filter captures cells while allowing medium to pass through, enabling concentration without direct mechanical stress on cells. This intermediary mechanism facilitates cell separation and concentration while preserving cell integrity and viability
2Reliability
If lower density stocks are used for cryopreservation, then cell viability is maintained, but productivity deteriorates due to inefficient inoculation of large volume cultures
Solution Approach 1:
The patent changes the cell density parameter during the cryopreservation process by implementing a two-stage approach: first maintaining cells at lower density during culture to preserve viability, then using the perfusion system to concentrate cells to high density before freezing. This parameter transformation allows the final cell bank to have both high density (improving inoculation efficiency) and high viability (from the gentle concentration process)
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
This approach allows for the creation of high-density frozen cell banks with excellent post-thaw viability, enabling efficient production cell culture and therapeutically relevant protein production.
Implementation Method 1
non-centrifugal cell retention systems, specifically alternating tangential flow filtration
Implementation Method 2
perfusion culture techniques coupled with non-centrifugal cell retention systems
Implementation Method 3
cryopreservation with DMSO addition
Implementation Method 4
adding DMSO to the concentrated cell population at a final concentration of about 5% to about 10%, vol/vol
Data Source
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AI summary
The current disclosure provides a method for the creation of a high-density cryopreserved cell bank using perfusion culture techniques and non-centrifugal concentration of cells. Methods of production using this high-density cryopreserved cell bank are also provided.