Spatial Capture Probe Arrays for Single-Cell Location Mapping
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Solution Overview
Problem
Existing methods fail to provide spatial information on the position of single cells within a biological sample, such as tissues, while capturing analyte data for a small handful of analytes or providing extensive data for individual cells without positional context.
Innovation Solution
The method involves generating a spatial array by attaching oligonucleotides to a substrate, extending primers, creating 3′ overhangs, and ligating second oligonucleotides with capture domains, which includes spatial barcodes, to capture and determine the location of analytes in a biological sample.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Loss of information
If traditional methods are used to capture analytes, then analyte data can be obtained for a small handful of analytes or for individual cells, but spatial information regarding the position of cells within the tissue is lost
Solution Approach 1:
The patent implements a multi-component oligonucleotide structure where a first oligonucleotide (attached to substrate) and a second oligonucleotide (containing capture domain) are ligated together. This nested structure allows the capture domain to remain accessible for analyte binding while the spatial barcode on the first oligonucleotide records positional information, thereby preserving both spatial context and analyte capture capability simultaneously.
Solution Approach 2:
The first oligonucleotide acts as an intermediary between the substrate and the capture domain. It contains a spatial barcode that records positional information and is ligated to the second oligonucleotide containing the capture domain. This intermediary structure enables the transfer of spatial information from the substrate location to the captured analyte without compromising the capture function.
2Measurement precision
If capture probes are designed to capture specific gene analytes, then gene-specific data is obtained, but the method becomes complex and requires multiple probe designs
Solution Approach 1:
The second oligonucleotide contains a universal capture domain structure that can bind to various analytes. Gene-specificity is achieved not through different capture domain designs but through the sequence content of the analyte itself hybridizing to the capture domain, allowing a single universal probe design to capture multiple different analytes with high precision.
Solution Approach 2:
The patent utilizes changes in the hybridization conditions and sequence complementarity parameters to achieve gene-specific detection. The capture domain's ability to specifically bind target analytes is controlled by the nucleic acid hybridization parameters (temperature, salt concentration, sequence match), simplifying the probe design while maintaining measurement precision.
3Quantity of substance
If extensive analyte data is provided for individual cells, then detailed molecular information is obtained, but information regarding the position of cells in the parent tissue sample is not provided
Solution Approach 1:
The method segments the information capture into two distinct parts: the first oligonucleotide captures spatial information through its position on the substrate and associated barcode, while the second oligonucleotide captures analyte information through its capture domain. This segmentation allows both spatial precision and extensive analyte data collection to occur independently and simultaneously.
Solution Approach 2:
The patent adds a spatial dimension to the traditional analyte capture by attaching oligonucleotides to a spatially-resolved substrate. The first oligonucleotide's position on the substrate (x, y coordinates) provides spatial information in addition to the molecular information captured by the second oligonucleotide, effectively adding a spatial dimension to the data output.
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 the precise determination of analyte locations within a biological sample, providing spatially resolved data on gene expression and protein distribution, enhancing understanding of cellular morphology and function.
Implementation Method 1
providing a splint oligonucleotide that hybridizes to the 3′ end of the first oligonucleotide
Implementation Method 2
extending the primer using the oligonucleotide as a template, thereby generating a first oligonucleotide with a free 3′ end
Implementation Method 3
ligating a second oligonucleotide to the 3′ end of the first oligonucleotide
Data Source
AI summary
The present disclosure relates to compositions and methods for generating capture probes on a substrate for identifying the location of analytes in a biological sample.