Sequential Hybridization Error Correction for Multiplex Imaging
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Solution Overview
Problem
Current methods for transcription profiling in cells, such as microscopy imaging, face challenges in accurately and efficiently identifying and quantifying multiple mRNAs in single cells, leading to errors and limitations in understanding cellular identity and disease treatment development.
Innovation Solution
A sequential hybridization method involving multiple rounds of hybridization with binding probes that emit detectable visual signals, allowing for error correction and unique code generation for each target gene, enabling efficient error reduction and accurate representation of gene expression levels.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If multiple mRNAs are identified and quantified in single cells using microscopy imaging, then valuable information regarding transcript abundance and localization is obtained, but errors occur in accurately identifying and quantifying multiple mRNAs
Solution Approach 1:
The patent divides the identification process into multiple sequential hybridization rounds, where each round targets specific subsets of mRNAs with unique barcode sequences. This segmentation allows systematic error correction by comparing results across rounds, thereby improving measurement precision while maintaining reliability.
Solution Approach 2:
The patent implements error correction mechanisms that use feedback from multiple hybridization rounds to identify and correct erroneous detections. By comparing barcode sequences across rounds and applying correction algorithms, the system improves the reliability of mRNA identification without sacrificing measurement precision.
2Measurement precision
If sequential hybridization with multiple rounds is performed to reduce errors, then accuracy of gene expression profiling is improved, but time and complexity of the process increase
Solution Approach 1:
The patent performs preliminary actions by designing and preparing multiple sets of binding probes with unique barcode sequences before the actual hybridization process. This preparation allows the sequential rounds to proceed efficiently with minimal optimization time, reducing the overall time loss while maintaining high measurement precision.
Solution Approach 2:
The patent optimizes hybridization parameters such as temperature, buffer composition, and probe concentration to reduce the time required for each round while maintaining detection accuracy. By carefully adjusting these parameters, the system achieves better measurement precision without excessive time investment.
3Adaptability or versatility
If binding probes emit detectable visual signals for each target gene, then identification of multiple mRNAs is enabled, but signal detection and differentiation becomes more difficult
Solution Approach 1:
The patent uses unique barcode sequences as intermediaries between the binding probes and the detection system. Each mRNA target is associated with a specific barcode, and detection is performed by reading these barcodes rather than directly detecting diverse visual signals. This intermediary approach enables versatile multi-target detection while simplifying the detection process.
Solution Approach 2:
The patent creates multiple copies of binding probes with identical barcode sequences that target the same mRNA. These replicated probes enhance the visual signal intensity for each target, making detection easier while maintaining the ability to distinguish multiple different targets through their unique barcodes.
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 method significantly reduces errors and enhances the accuracy of gene expression profiling, allowing for precise identification and quantification of multiple mRNAs in single cells, thereby improving our understanding of cellular identity and disease treatment development.
Implementation Method 1
sequential hybridization of binding probes to the plurality of target genes
Implementation Method 2
a binding sequence that specifically binds a target sequence in a gene
Implementation Method 3
each probe is capable of emitting a detectable visual signal upon binding of the probe to a target sequence
Data Source
AI summary
Disclosed herein are methods and systems for detecting and/or quantifying cellular targets such as nucleic acids in cells, tissues, organs or organisms. Through sequential barcoding, it is possible to perform high-throughput profiling of a large number of targets, such as transcripts and/or DNA loci. In some embodiments, error correction is implemented through use of barcodes that can tolerate mistakes and missing data during sequential hybridization of probes to selected targets.