Expansion Sequencing Spatial Resolution in Biological Samples
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Solution Overview
Problem
Current methods for analyzing biomolecules like DNA and RNA in tissues lack the ability to simultaneously maintain spatial location and multiplexed measurement, with optical methods limited in the number of molecules that can be studied and transcriptomic approaches losing spatial information, particularly in brain tissues where neuronal communication details are crucial.
Innovation Solution
Expansion sequencing (ExSEQ) method, which leverages expansion microscopy (ExM) to physically expand biological samples, allowing for in situ nucleic acid sequencing with techniques like FISSEQ, enabling spatially-localized, long-read sequencing and validation of sequencing data through alignment with in vitro sequencing technologies.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Loss of information
If optical methods are used to maintain spatial location of biomolecules, then spatial information is preserved, but the number of biomolecules that can be studied simultaneously is limited
Solution Approach 1:
The patent applies expansion microscopy to physically expand the biological sample in three-dimensional space, creating additional spatial dimensions. This expansion allows optical microscopy to resolve and simultaneously study a much larger number of biomolecules while preserving their spatial relationships, effectively transitioning from a limited 2D projection to an expanded 3D visualization space
2Quantity of substance
If transcriptomic approaches are used to allow multiplexed measurement of all RNA and DNA molecules, then the number of biomolecules studied increases, but spatial information is lost
Solution Approach 1:
The patent performs preliminary expansion of the biological sample before conducting transcriptomic analysis. By expanding the sample structure in advance, the spatial coordinates of all RNA and DNA molecules are preserved and magnified, allowing subsequent multiplexed sequencing to maintain spatial information that would otherwise be lost in conventional transcriptomic approaches
3Measurement precision
If in situ sequencing is performed on expanded samples, then spatial resolution is improved, but the length of sequencing reads is currently limited
Solution Approach 1:
The patent employs dynamic oligonucleotide extension strategies where sequencing reads are progressively extended through multiple iterative cycles. The system dynamically adjusts extension conditions and uses polymerase chain reaction or rolling circle amplification to lengthen reads in situ, allowing the sequencing read length to adaptively increase while maintaining the spatial resolution benefits of expansion microscopy
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 systematic cell type and cell state classification with nanoprecise spatial resolution, preserving spatial information and allowing for combinatorial measurements of epigenomic changes, enhancing understanding of cellular and organ-scale functions and pathologies.
Implementation Method 1
the composition can be expanded isotropically, preferably with nanoscale precision, in three dimensions
Data Source
AI summary
The invention provides in situ nucleic acid sequencing to be conducted in biological specimens that have been physically expanded. The invention leverages the techniques for expansion microscopy (ExM) to provide new methods for in situ sequencing of nucleic acids in a process referred to herein as “expansion sequencing” (ExSEQ).