Microfluidic Spatial Encoding for High-Multiplex Biological Assays
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Solution Overview
Problem
Existing methods for analyzing spatial expression patterns of biological molecules, such as genes and proteins, are limited in their ability to simultaneously measure multiple targets at high resolution and scale, and lack reproducibility.
Innovation Solution
A method using a microfluidic device with multiple addressing channels to deliver probes and address tags at intersecting angles, allowing for the determination of spatial patterns of biological target abundance, expression, and activity by analyzing probes bound to the target.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional methods such as in situ hybridization or microarrays are used, then spatial patterns of gene expression can be analyzed, but the ability to simultaneously measure multiple targets at high resolution is limited
Solution Approach 1:
The patent introduces a combinatorial addressing system that adds a second spatial dimension (X and Y coordinates) to the measurement process. By delivering probes along two perpendicular sets of addressing channels intersecting at right angles, the system achieves high spatial resolution while simultaneously measuring thousands of genetic targets through the combination of address tags from both dimensions.
Solution Approach 2:
The patent segments the measurement process into multiple independent components: probes with specific binding moieties for different targets, address tags for spatial location, and combining groups that link targets to locations. This segmentation allows each component to be optimized independently while achieving high multiplexing capability and spatial resolution simultaneously.
2Measurement precision
If laser capture microdissection is used to analyze many genes at small locations, then spatial information is preserved, but the method is very expensive, laborious, and does not scale well
Solution Approach 1:
The patent replaces the manual, mechanical laser capture microdissection process with an automated microfluidic system. Addressing channels and probes are delivered through fluidic pathways, and spatial information is encoded through combinatorial address tags rather than physical manipulation, enabling high-throughput automated analysis that scales efficiently.
Solution Approach 2:
The patent changes the fundamental parameters of the analysis system by using microfluidic flow rates, probe concentrations, and addressing channel geometries that enable thousands of simultaneous measurements. This parameter optimization allows the system to achieve high productivity while maintaining spatial precision, unlike the fixed-capacity laser capture method.
3Measurement precision
If PCR assays in a 2D format are used, then spatial information is preserved, but spatial resolution is low because they rely on physically transferring tissues into wells
Solution Approach 1:
The patent transitions from a single-plane 2D well array to a true two-dimensional addressing system with intersecting X and Y addressing channels. This dimensional enhancement allows probes to be delivered precisely to any intersection point, achieving high spatial resolution while maintaining the ability to preserve spatial information through combinatorial address tag encoding.
4Adaptability or versatility
If high levels of multiplexing are achieved, then many targets can be measured simultaneously, but random access to tissue samples is prevented
Solution Approach 1:
The patent performs preliminary actions by pre-positioning addressing channels and probes before the actual measurement process. The combinatorial addressing system is established in advance, allowing any location to be accessed by delivering the appropriate combination of address tags through the pre-configured channel network, thus maintaining random access capability while achieving high multiplexing.
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 high-resolution, reproducible spatial mapping of large numbers of biological molecules simultaneously, with the potential to analyze hundreds of thousands to millions of probes in parallel.
Implementation Method 1
delivering a probe for a biological target to a sample... delivering a first address tag through each of the first addressing channels to each first area in the sample
Implementation Method 2
a binding moiety capable of binding to the biological target
Data Source
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AI summary
The present disclosure provides methods and assay systems for use in spatially encoded biological assays, including assays to determine a spatial pattern of abundance, expression, and/or activity of one or more biological targets across multiple sites in a sample. In particular, the present disclosure provides methods and assay systems capable of high levels of multiplexing where reagents are provided to a biological sample in order to address tag the sites to which reagents are delivered; instrumentation capable of controlled delivery of reagents, in particular, microfluidic device based instrumentation; and a decoding scheme providing a readout that is digital in nature.