Droplet Microfluidic Neutralization Assays for ASC Screening

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Solution Overview

Problem

Current methods for screening antibody-secreting cells (ASCs) are limited by low throughput and inability to perform functional neutralization assays, leading to the overlook of promising candidates and potential Antibody-Dependent Enhancement effects.

Innovation Solution

A high-throughput droplet microfluidic system for isolating and retrieving ASCs based on neutralizing function, involving droplet encapsulation, picoinjection of virus and host cells, and dielectrophoretic sorting to enrich for cells secreting neutralizing antibodies.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Productivity

If FACS and display systems are used to screen ASCs for binding to protein antigen, then large numbers of cells can be screened, but binding affinity is not reflective of true functional efficacy and non-neutralizing epitopes are selected

Engineering Contradiction:

Improvescreening throughputVSAvoidfunctional efficacy measurement

Core Design Contradiction:

ProductivityVSMeasurement precision

Solution Approach 1:

The patent uses an intermediary virus neutralization assay system that bridges the gap between binding affinity measurement and functional efficacy. The system employs droplet-based microfluidics to create isolated reaction environments where virus-antibody-neutralizing antibody interactions can be measured directly, providing an intermediary measurement that reflects true functional efficacy while maintaining high throughput screening capability

Inventive Principle:

Principle #24Intermediary (Mediator)

Solution Approach 2:

The patent replaces the mechanical FACS sorting system with a microfluidic droplet-based system that uses flow dynamics and droplet manipulation to achieve high-throughput screening. This substitution allows for integrated functional assays within the screening workflow, enabling direct measurement of virus neutralization activity rather than relying on surrogate binding measurements

Inventive Principle:

Principle #28Mechanics substitution (Replace mechanical system)

2Measurement precision

If hybridoma generation and single B cell activation/expansion are used to screen for effective nAbs, then functional neutralization can be assessed, but the laborious nature results in long workflow and low throughput

Engineering Contradiction:

Improvefunctional neutralization assessmentVSAvoidscreening throughput

Core Design Contradiction:

Measurement precisionVSProductivity

Solution Approach 1:

The patent segments the traditional sequential hybridoma workflow into parallel microfluidic operations. By encapsulating individual B cells in droplets and performing neutralization assays directly within droplets, the system eliminates time-consuming cell culture and expansion steps, enabling simultaneous processing of thousands of candidates while maintaining functional assessment accuracy

Inventive Principle:

Principle #1Segmentation

Solution Approach 2:

The patent performs preliminary action by pre-encapsulating B cells in droplets with necessary assay reagents before the actual neutralization assay. This preliminary preparation allows direct functional testing without requiring prior cell expansion or activation, dramatically reducing workflow time while maintaining the ability to assess functional neutralization

Inventive Principle:

Principle #10Preliminary action

3Measurement precision

If conventional virus neutralization assays are used, then functional neutralization can be measured, but the low throughput limits the immune repertoire that can be evaluated

Engineering Contradiction:

Improvevirus neutralization measurementVSAvoidnumber of candidates screened

Core Design Contradiction:

Measurement precisionVSProductivity

Solution Approach 1:

The patent applies pneumatic and hydraulic principles through microfluidic droplet generation and manipulation. By using flow-focusing geometry and pressure-driven flow to create and control droplets, the system achieves high-throughput processing of virus neutralization assays, enabling evaluation of large numbers of candidates while maintaining precise functional measurement in each droplet

Inventive Principle:

Principle #29Pneumatics and hydraulics

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

Enables rapid discovery of potent neutralizing antibody-producing cells, achieving high-throughput screening and enrichment of functional ASCs, overcoming limitations of existing low-throughput and non-functional screening methods.

Implementation Method 1

droplet sorting based on neutralizing activity

Methodology Applied

Scientific EffectDielectrophoresis: Dielectric Permittivity

Data Source

PatentUS20250353012A1Microfluidic assays and uses thereof

Publication Date: 2025.11.20 NATIONAL UNIVERSITY OF SINGAPORE

AI summary

The present disclosure concerns a microfluidic method of assaying antibody secreting cells (ASCs), comprising the steps of isolating ASCs within droplets such that each droplet encapsulates only one ASC; incubating the droplets of step a) to accumulate antibodies within the droplets; picoinjecting virus into the droplets of step b) to form immune complex droplets; picoinjecting host cells into the immune complex droplets to form neutralised droplets and infected droplets; and sorting the infected droplets from the neutralised droplets, b based on infection of the host cells by the virus, to assay the ASCs within the neutralised droplets. The present disclosure also concerns a microfluidic platform thereof.