Microfluidic Droplet Assays for Single-Cell Effector Screening
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Solution Overview
Problem
Current methods for selecting and growing effector cells, such as those used in adoptive cell therapy, are limited by the need for large volumes of cytokine secretion assays and the inability to effectively identify individual effector cells specific to disease-related antigens.
Innovation Solution
The method involves encapsulating an immune cell, a target cell, a first signaling entity for determining cell viability, and a second signaling entity for determining cytokine secretion within a microfluidic droplet, allowing for the detection and sorting of droplets containing active effector cells.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If limiting dilution method is used in microwell plates, then cell selection can be performed, but cytokine secretion from single cells cannot be detected due to large minimum working volume
Solution Approach 1:
The patent segments the assay into individual microfluidic droplets, each containing a single cell. This segmentation allows cytokine secretion to be detected at single-cell resolution, overcoming the limitation of bulk microwell plate assays where cytokines from multiple cells are diluted and difficult to detect. Each droplet acts as an independent measurement unit, enabling sensitive detection of cytokines secreted by individual effector cells.
2Measurement precision
If well-based cytokine secretion assay is used, then cytokine detection can be performed, but hundreds of cells per well are required which prevents association of single effector cells with specific target antigens
Solution Approach 1:
The patent divides the population of effector cells into individual droplets, each containing a single cell. This segmentation enables tracking and association of specific effector cells with their target antigen presentations. By isolating cells into separate droplets, the system can determine which specific effector cell interacts with which specific target cell, providing single-cell resolution that is impossible in bulk well-based assays.
Solution Approach 2:
The patent introduces target cells as intermediaries that present disease-related antigens to effector cells within the droplet. This intermediary mechanism enables specific antigen-cell interactions to be captured and analyzed, allowing the system to identify which effector cells recognize specific antigens by observing interactions between the intermediary target cells and the effector cells in the same droplet.
3Productivity
If limiting dilution method is used, then effector cell selection can be performed, but throughput is low and many generations of cell expansion are required
Solution Approach 1:
The patent performs preliminary identification and selection of functional effector cells at the single-cell level using high-throughput droplet-based cytokine secretion assays. By screening and identifying active effector cells early in the process, the system can reduce or eliminate the need for multiple generations of cell expansion, as the selected effector cells are already characterized for their functional activity against specific targets.
Solution Approach 2:
The patent replaces the mechanical, labor-intensive limiting dilution method with an automated microfluidic droplet-based system. This substitution enables parallel processing of thousands of single-cell assays simultaneously, dramatically increasing throughput. The automated detection and sorting of droplets containing active effector cells eliminates manual intervention and accelerates the identification process, reducing the time required for cell expansion and therapy production.
Data Source
AI summary
The present invention generally relates to fluidic droplets and systems and methods for determining immune or other cells. Some aspects of the invention are generally directed to assays that combine sensitive detection of secreted products with detection of target cell death in droplets containing an effector cell, systems and methods to isolate droplets in which one or more cell interactions have occurred, or systems and methods to generate nucleic acid information from cell interactions. In addition, some embodiments of the invention are generally directed to containing two (or more) cells in droplets, e.g., an effector cell and one or more target cells, and determining various interactions between the cells within the droplets, such as whether the effector cell kills the target cell, whether the effector cell releases antibodies, cytokines or other substances that are able to interact with the target cell or are released in the presence of the target cell, or the like.