Peripheral Capture Probes to Reduce Analyte Mislocalization
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Solution Overview
Problem
Spatial analysis assays in biological samples suffer from mislocalization of analytes, leading to an artificially elevated 'hot edge' signal pattern at the perimeter of the array, which distorts detection signals and reduces the accuracy of spatial analysis.
Innovation Solution
Incorporating capture probes at the periphery of the array that do not include primer binding sites or sequencing specific sites, thereby capturing analytes that are not ultimately detected, to mitigate the mislocalization and hot edge signal pattern.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If the biological sample extends beyond the boundaries of the capture probes on the array, then more analytes can be captured, but mislocalization occurs leading to hot edge signal pattern and reduced measurement precision
Solution Approach 1:
The array is segmented into two distinct types of capture probes: detecting capture probes (with spatial barcodes and functional domains) and non-detecting capture probes (without spatial barcodes or functional domains). This segmentation allows the system to differentiate between analytes that should be spatially localized and those that should be absorbed to prevent mislocalization, thereby resolving the contradiction between capturing more analytes and maintaining spatial analysis accuracy.
Solution Approach 2:
Non-detecting capture probes act as intermediary elements at the periphery of the array. These probes serve as a buffer zone that captures migrating analytes before they can be mislocalized to detecting capture probes. By introducing this intermediary layer, the system maintains high analyte capture capacity while preventing spatial distortion and hot edge artifacts.
2Reliability
If capture probes are placed at the periphery of the array, then analyte migration can be captured, but artificial signal elevation occurs at the perimeter
Solution Approach 1:
Instead of making all peripheral capture probes detecting probes (which would cause hot edge artifacts), the invention inverts the approach by making peripheral probes non-detecting. This inversion allows the periphery to fulfill its analytical function of capturing migrating analytes without generating false positive signals, thus maintaining both capture completeness and signal accuracy.
Solution Approach 2:
Different regions of the array are assigned different functional qualities: central regions contain detecting capture probes with full functionality for spatially resolved detection, while peripheral regions contain non-detecting capture probes optimized for capturing migrating analytes. This local differentiation ensures that each region performs its specific function optimally without interfering with other regions, resolving the contradiction between capture completeness and signal accuracy.
3Productivity
If all capture probes include primer binding sites and sequencing specific sites, then all captured analytes can be detected, but mislocalized analytes generate false signals
Solution Approach 1:
The capture probe population is segmented into two functional categories: detecting capture probes that include spatial barcodes and functional domains for sequencing and detection, and non-detecting capture probes that lack these elements. This segmentation enables the system to process and detect analytes efficiently while preventing mislocalized analytes from generating false spatial signals, as non-detecting probes cannot produce detectable output even when they capture analytes.
Solution Approach 2:
The invention changes the functional parameters of peripheral capture probes by removing primer binding sites and sequencing specific sites. This parameter modification prevents these probes from participating in the detection and sequencing process, thereby eliminating their contribution to false spatial signals while maintaining their ability to capture migrating analytes and improve overall detection throughput.
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
Improves the accuracy of spatial transcriptomics assays by reducing mislocalization of analytes, ensuring accurate spatially derived data without the artificial elevation of signals at the array's periphery.
Implementation Method 1
hybridizing target nucleic acids from the biological sample to the first capture domain of capture probes of the first region and a second capture domain of the capture probes of the second region
Data Source
AI summary
Compositions, kits, and methods for reducing mislocalization of analytes from a biological sample in the context of an array-based spatial analysis platform are disclosed herein. Also, disclosed herein are a first region of capture probes, where the capture probes include: (i) a spatial barcode, (ii) a first capture domain, and (iii) one or more functional domains, and a second region of capture probes, where the capture probes include a second capture domain. The second region of capture probes can capture analytes from portions of the biological sample that exceed the boundaries of the first region of capture probes, thereby reducing analyte mislocalization and improving the accuracy of the array-based spatial analysis platform.