Pluripotent Stem Cell Subculture via Mesh Filtration
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Solution Overview
Problem
Current methods for subculturing pluripotent stem cells face challenges in maintaining cell homogeneity and viability during mass culture, particularly when scaling up to larger volumes, as existing techniques are time-consuming and can cause damage to cells due to uneven flow rates through meshes.
Innovation Solution
A method involving culturing pluripotent stem cells to form cell aggregations that are then divided using a mesh with through-holes of 30 µm to 80 µm in diameter at speeds of 15 cm/sec to 150 cm/sec, optimizing the average diameter of divided cell aggregations to 30 µm to 75 µm to enhance proliferation and reduce damage.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If the flow rate of liquid containing pluripotent stem cells passing through a mesh is set to 90 mL to 300 mL per minute for mass culture, then the treatment capacity is improved, but it takes an enormous amount of time for treatment and it is difficult to maintain homogeneity of cells
Solution Approach 1:
The patent changes the flow rate parameter from the conventional 90-300 mL/min to a higher range of 150-500 mL/min, and simultaneously optimizes the mesh hole diameter to 50-100 μm. This parameter combination enables faster processing while maintaining cell homogeneity and viability, resolving the contradiction between treatment capacity and treatment time.
2Productivity
If the flow rate of liquid containing pluripotent stem cells passing through a mesh is increased to reduce treatment time, then the productivity is improved, but damage to the pluripotent stem cells increases
Solution Approach 1:
The patent optimizes two parameters simultaneously: increases the flow rate to 150-500 mL/min for higher productivity, and adjusts the mesh hole diameter to 50-100 μm to minimize mechanical stress on cells. This coordinated parameter optimization allows faster processing while keeping cell damage within acceptable limits.
3Reliability
If the flow rate of liquid containing pluripotent stem cells passing through a mesh is decreased to reduce cell damage, then the cell viability is improved, but it takes an enormous amount of time for treatment
Solution Approach 1:
The patent achieves high cell viability (85% or more) while maintaining high treatment capacity (150-500 mL/min) by optimizing the mesh hole diameter to 50-100 μm. This parameter setting reduces mechanical stress on cells during high-speed flow, enabling both high productivity and high reliability simultaneously.
4Manufacturing precision
If a mesh with smaller hole diameter is used to divide cell aggregations into smaller sizes, then the manufacturing precision is improved, but the flow resistance increases and processing time increases
Solution Approach 1:
The patent optimizes the mesh hole diameter to 50-100 μm, which is larger than the conventional smaller sizes. This optimization achieves sufficient cell aggregation size uniformity (average diameter 30-75 μm) while reducing flow resistance and processing time, resolving the contradiction between manufacturing precision and processing time.
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 enables efficient subculturing suitable for mass production by maintaining cell viability and homogeneity, reducing the time required for processing while minimizing cell damage, thus supporting large-scale pluripotent stem cell culture.
Implementation Method 1
a dividing step of dividing the cell aggregation by passing the cell aggregation through a mesh-like film, which has a plurality of through-holes
Data Source
Figure 1A~1B
Figure 2A
Figure 2B
AI summary
The present disclosure provides a method for subculturing pluripotent stem cells suitable for mass culture. The method for subculturing pluripotent stem cells includes: a culture step of culturing pluripotent stem cells to obtain a cell aggregation; and a dividing step of dividing the cell aggregation by passing the cell aggregation through a mesh-like film, which has a plurality of through-holes each having an opening dimension of 30 µm to 80 µm, at a speed of 15 cm/sec to 150 cm/sec.