Microfluidic Droplet Screening for High-Complexity Molecular Interactions
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Solution Overview
Problem
Current protein-protein interaction screening methods are limited by small library sizes and low throughput, particularly in identifying complex interactions between molecules, as they often require constant interaction partner conditions and sequential sequencing of individual clones, which restricts the efficiency and complexity of screening.
Innovation Solution
A method involving two separated libraries of candidate interacting entities, where each entity comprises an interacting-portion and a labelling-portion, allowing for the formation of interaction-complexes in microfluidic compartments at low entity concentrations, enabling the detection of interacting pairs through fusion PCR and nucleic acid-based hybridization techniques.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If classical bait and prey screening methods are used, then interaction identification is achieved, but library size is limited and throughput is low
Solution Approach 1:
The invention segments the screening process by encapsulating individual bait-prey interaction pairs in separate microfluidic droplets. This segmentation enables parallel processing of numerous interaction pairs simultaneously, thereby increasing throughput while accommodating large library sizes. Each droplet acts as an independent reaction chamber, allowing high-complexity screening without compromising interaction detection accuracy.
2Productivity
If constant interaction partner conditions are maintained, then reliable interaction detection is achieved, but screening complexity and efficiency are reduced
Solution Approach 1:
The invention applies local quality by maintaining constant interaction partner conditions only within each individual droplet, while allowing variation across different droplets to accommodate diverse library members. This localized control ensures reliable interaction detection for each pair while enabling high-throughput screening of complex libraries with diverse interaction partners.
3Loss of time
If sequential sequencing of individual clones is performed, then interaction pairs are identified, but time consumption increases and throughput decreases
Solution Approach 1:
The invention merges the identification of interaction pairs by performing PCR amplification and sequencing on pooled droplets containing multiple interaction pairs. This combining approach eliminates the need for sequential processing of individual clones, dramatically reducing time consumption while maintaining identification accuracy through the use of unique molecular identifiers that allow computational deconvolution of pooled sequences.
4Quantity of substance
If high concentration of entities is used, then interaction formation is promoted, but false positive interactions increase
Solution Approach 1:
The invention extracts the interaction detection from the bulk solution by encapsulating individual interaction pairs in separate droplets. This extraction allows the use of sufficiently high entity concentrations within each droplet to promote interaction formation, while the physical separation prevents false positive interactions between unrelated entities in different droplets. Each droplet effectively isolates its own interaction chemistry.
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 the identification of interacting entities with higher complexity and efficiency, allowing for the screening of larger candidate libraries and specific immunological interactions, such as T-cell receptor and antigen presenting cell interactions, by encapsulating interaction-complexes in microfluidic compartments at low entity concentrations and using PCR to amplify and identify interacting sequences.
Implementation Method 1
using PCR to amplify and identify interacting sequences
Implementation Method 2
using nucleic acid-based hybridization techniques
Data Source
AI summary
The invention regards a method for screening for interactions between two compounds provided in libraries. The invention in particular provides a method for screening interactions, such as a binding between two molecules or macromolecules, that is not limited to a classical screening by bait and prey setup, and therefore allows for a high-complexity screening of large candidate libraries, including screening for immunological interactions such as interactions between T-cells and antigen presenting cells, or B-cells and their antigenic targets.