Microfluidic In-Pen Assays for Position-Insensitive Analyte Quantification
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Solution Overview
Problem
Existing technologies face challenges in accurately determining the quantity of analytes produced by micro-objects confined within microfluidic chambers, particularly due to sensitivity to the position of biological micro-objects and limitations in measuring analyte concentration fluctuations.
Innovation Solution
A system and method involving an image acquisition unit with an image processing unit to define an area of interest within sequestration pens in a microfluidic device, analyzing image areas to determine analyte quantity, using reporter molecules to detect analyte-secreted complexes, and selecting top analyte-producing micro-objects for expansion.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If the entire sequestration pen area is used for analyte measurement, then the measurement area is maximized, but the position sensitivity to biological micro-objects increases
Solution Approach 1:
The sequestration pen area is segmented into multiple sub-regions, with at least one sub-region designated as the area of interest for analyte measurement. This segmentation allows the system to exclude regions containing biological micro-objects from the measurement area, thereby reducing position sensitivity while maintaining adequate measurement area for accurate analyte concentration detection.
2Object-affected harmful factors
If the area of interest is reduced to minimize position sensitivity, then position sensitivity decreases, but the measurement area for analyte detection is reduced
Solution Approach 1:
Different regions of the sequestration pen are assigned different functional qualities: the area of interest is optimized for analyte measurement with specific optical properties, while other regions may contain biological micro-objects or serve as control areas. This local differentiation allows the system to maintain measurement accuracy in the designated area while accommodating biological micro-objects in other regions.
3Loss of information
If image processing is performed on the entire sequestration pen, then comprehensive data is obtained, but the computational complexity and processing time increase
Solution Approach 1:
The image processing system extracts and focuses computational resources on the area of interest within the sequestration pen, rather than processing the entire pen area. This extraction approach maintains comprehensive data for the measurement region while significantly reducing computational complexity and processing time by excluding regions containing biological micro-objects or non-measurement areas.
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 precise quantification of analyte production by micro-objects, minimizing position sensitivity and enhancing the selection of high-producing micro-objects for further analysis.
Implementation Method 1
allowing a portion of the plurality of reporter molecules to diffuse into the sequestration pen and bind to the analyte secreted therein
Implementation Method 2
each reporter molecule includes: a binding component configured to bind the secreted analyte
Implementation Method 3
an imaging element configured to capture one or more assay images of the plurality of sequestration pens and the flow region
Data Source
AI summary
Methods, systems and kits are described herein for detecting the results of an assay. In particular, the methods, systems and devices of the present disclosure rely on a difference between the diffusion rates of a reporter molecule and an analyte of interest in order to quantify an amount of analyte in a microfluidic device. The analyte may be a secreted product of a biological micro-object.