Continuous Non-Viral Transfection via Passive Mixing Fluidic Module
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Solution Overview
Problem
Current methods for producing non-viral carrier nucleic acid particles for cell transfection are discontinuous and prone to variability, making them less reliable and challenging for large-scale production due to the need for incubation times that hinder continuous processing.
Innovation Solution
A method involving a passive mixing fluidic module with separate inlets for nucleic acid and transfection agent solutions, which are combined and flowed through a cell reactor with controlled flow rates and residency times, allowing for continuous and scalable production of transfection complexes.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Ease of manufacture
If bulk mixing method is used to prepare DNA-transfection agent complexes, then the process is simple to implement, but the production is discontinuous and batch-to-batch variability is high
Solution Approach 1:
The patent implements continuous transfection by continuously adding transfection agent to nucleic acid in a flow-through system, eliminating the discontinuous bulk mixing approach. The transfection agent solution is continuously pumped through a needle valve into the nucleic acid solution, maintaining continuous complex formation and delivery to cells, thereby improving batch-to-batch reproducibility while retaining operational simplicity.
2Stability of the object's composition
If incubation time of 5 to 30 minutes is provided for complex formation, then stable DNA-transfection agent complexes can form, but continuous production is prevented
Solution Approach 1:
The patent achieves continuous complex formation by using a flow-through system where transfection agent is continuously added to nucleic acid. The continuous pumping action via needle valve maintains steady-state complex formation without requiring discrete incubation periods, thereby enabling continuous production while maintaining complex stability through controlled residence time in the flow path.
Solution Approach 2:
The patent changes the temporal parameter from discrete incubation intervals to continuous flow residence time. By controlling the flow rate and residence time of the transfection agent solution in the needle valve and tubing system, the patent achieves continuous complex formation that replaces the traditional 5-30 minute incubation step, enabling uninterrupted production.
3Quantity of substance
If manual mixing of large volumes is performed, then bulk mixing can be achieved, but handling difficulty increases
Solution Approach 1:
The patent replaces manual mechanical mixing with a flow-through needle valve system that uses fluid dynamics to achieve mixing. The needle valve controls the flow rate and mixing efficiency without requiring manual intervention, thereby enabling handling of large volumes with automated control and eliminating the difficulty of manual mixing operations.
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 reproducible and continuous transfection of cells, reducing batch-to-batch variability and facilitating large-scale production of recombinant proteins and viral particles by maintaining consistent transfection complex formation and uptake.
Implementation Method 1
providing the NAS at a first flow rate; providing the TAS at a second flow rate; providing a combined stream of the NAS and TAS from the single outlet to the cell reactor through a length of tubing
Data Source
AI summary
Methods and systems are provided for transfecting cells using real-time, continuous transfection of cells. In some aspects, the methods can be applied for the continuous production of non-viral vector nucleic acid complexes. The systems and methods include a passive mixing fluidic module with at least two inlets, a plurality of mixing elements, and an outlet to provide a continuous flow of transfection complexes to a cell reactor. The transfection agent and nucleic acid are passively mixed and then provided to cells in a continuous flow of cell medium. In some aspects, the flow of cell medium perfusing through the cell reactor recirculates. The system and the methods of the present disclosure provide for highly reproducible and scalable transfection with a low coefficient of variation.