Affinity-Based Partition Assay for Biomolecule Detection
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Solution Overview
Problem
Current methods for detecting and quantifying biomolecules, such as ELISA and immunoPCR, face limitations in precision and sensitivity due to high background signals from non-specific binding of antibodies, which hinder accurate detection and quantification of biomolecules.
Innovation Solution
A method involving the use of two or more probes linked to distinct labels that specifically bind to a target molecule, with the sample partitioned into multiple partitions to enhance the likelihood of co-localization of probes with the target, allowing for improved detection and quantification by distinguishing specific binding from random co-localization.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If immunoPCR is used to increase detection sensitivity, then the sensitivity of detection is improved, but high background signal due to non-specific binding of antibody worsens the measurement precision
Solution Approach 1:
The sample is partitioned into many small partitions (droplets or wells), distributing probes and target molecules across multiple compartments. This segmentation allows individual partitions to contain at most one target molecule, enabling precise counting while reducing non-specific binding effects that plague bulk assays like immunoPCR.
Solution Approach 2:
Multiple probes specific to different epitopes of the same target molecule are used as intermediaries. The co-localization of multiple differently labeled probes in the same partition serves as a specific indicator of target presence, distinguishing true signals from non-specific background binding.
2Measurement precision
If multiple probes linked to distinct labels are used to improve target identification accuracy, then the precision of quantification is improved, but the device complexity increases
Solution Approach 1:
Multiple probes targeting different epitopes of the same antigen are employed, each labeled with a distinct label. This multi-functional approach allows simultaneous detection and quantification of the target molecule through co-localization of multiple probes in the same partition, enhancing precision without requiring separate assays.
Solution Approach 2:
The use of multiple probes with distinct labels creates redundant detection signals. By requiring co-localization of multiple labeled probes in the same partition, the system verifies target presence through multiple independent signals, improving quantification precision while managing complexity through standardized probe design.
3Measurement precision
If partitioning is performed to reduce background noise and enhance detection sensitivity, then the limit of detection is improved, but the loss of time in partitioning and analysis increases
Solution Approach 1:
The assay partitions the sample into numerous small compartments, enabling parallel processing of many samples simultaneously. This segmentation achieves high sensitivity by distributing target molecules across partitions, with each partition analyzed independently, reducing background noise while maintaining efficient throughput.
Solution Approach 2:
Probes are pre-labeled with distinct labels and prepared in advance. The partitioning step is optimized to distribute probes and targets efficiently, and signal detection is performed in parallel across all partitions, reducing the overall assay time despite the additional partitioning step.
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 enhances the sensitivity and precision of biomolecule detection and quantification by reducing background noise and increasing the accuracy of target molecule identification, enabling lower limits of detection and improved sensitivity.
Implementation Method 1
at least a first probe linked to a first label and a second probe linked to a second label, wherein the first and second probes specifically bind to the same target molecule
Data Source
Figure 1A~1F
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AI summary
The present invention provides methods of detecting a target molecule in a sample comprising incubating the sample with two or more detectably labeled probes, partitioning the sample into multiple partitions, and detecting the presence of the two or more probes in the same partition. Quantifying the amount of biomolecules in a sample from a subject can provide useful information for a number of clinical applications.