METHODS AND SYSTEMS FOR DETERMINING SPATIAL PATTERNS OF BIOLOGICAL TARGETS IN A SAMPLE
First Claim
1. A method of determining a spatial pattern of abundance, expression, and/or activity of one or more biological targets across multiple sites in a sample, comprising:
- delivering a probe for each of one or more biological targets to multiple sites in a sample, wherein each probe comprises;
(1) a target-binding moiety capable of binding to the probe'"'"'s corresponding biological target;
(2) an address tag that identifies each of the multiple sites to which the probe is delivered; and
(3) an identity tag that identifies the probe'"'"'s corresponding biological target or target-binding moiety;
allowing each probe to bind to its corresponding biological target in the sample;
analyzing the probe bound to the one or more biological targets, the analysis comprising;
(1) determining abundance, expression, and/or activity of each of the one or more biological targets by assessing the amount of the probe bound to the biological target; and
(2) determining the identities of the identity tag and the address tag of the probe; and
determining a spatial pattern of abundance and/or activity of the one or more biological targets across the multiple sites in the sample based on the analysis,wherein the method does not depend on an imaging technique for determining spatial information of the one or more biological targets in the sample.
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Accused Products
Abstract
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 biological targets comprise proteins, and the methods and assay systems do not depend on imaging techniques for the spatial information of the targets. 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; and a decoding scheme providing a readout that is digital in nature.
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Citations
64 Claims
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1. A method of determining a spatial pattern of abundance, expression, and/or activity of one or more biological targets across multiple sites in a sample, comprising:
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delivering a probe for each of one or more biological targets to multiple sites in a sample, wherein each probe comprises;
(1) a target-binding moiety capable of binding to the probe'"'"'s corresponding biological target;
(2) an address tag that identifies each of the multiple sites to which the probe is delivered; and
(3) an identity tag that identifies the probe'"'"'s corresponding biological target or target-binding moiety;allowing each probe to bind to its corresponding biological target in the sample; analyzing the probe bound to the one or more biological targets, the analysis comprising;
(1) determining abundance, expression, and/or activity of each of the one or more biological targets by assessing the amount of the probe bound to the biological target; and
(2) determining the identities of the identity tag and the address tag of the probe; anddetermining a spatial pattern of abundance and/or activity of the one or more biological targets across the multiple sites in the sample based on the analysis, wherein the method does not depend on an imaging technique for determining spatial information of the one or more biological targets in the sample. - View Dependent Claims (5, 6, 7, 8, 9, 10, 11, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51)
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2. A method of determining a spatial pattern of abundance, expression, and/or activity of one or more biological targets across multiple sites in a sample, comprising:
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delivering a probe for each of one or more biological targets to multiple sites in a sample, wherein each probe comprises;
(1) a target-binding moiety capable of binding to the probe'"'"'s corresponding biological target;
(2) an address tag having a sequence that identifies each of the multiple sites to which the probe is delivered; and
(3) an identity tag having a sequence that identifies the probe'"'"'s corresponding biological target or target-binding moiety;allowing each probe to bind to its corresponding biological target in the sample; analyzing the probe bound to the one or more biological targets by sequencing, wherein the amount of a sequencing product indicates abundance, expression, and/or activity of each of the one or more biological targets, the sequencing product comprising all or a portion of the address tag sequence and all or a portion of the identity tag sequence; and determining a spatial pattern of abundance, expression, and/or activity of the one or more biological targets across the multiple sites in the sample based on the analysis.
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3. A method of determining a spatial pattern of abundance, expression, and/or activity of one or more biological targets across multiple sites in a sample, comprising:
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delivering a probe for each of one or more biological targets to multiple sites in a sample, wherein each probe comprises;
(1) a target-binding moiety capable of binding to the probe'"'"'s corresponding biological target; and
(2) an identity tag that identifies the probe'"'"'s corresponding biological target or target-binding moiety;allowing each probe to bind to its corresponding biological target in the sample; delivering an address tag to each of the multiple sites in the sample, wherein the address tag is to be coupled to the probe bound to the biological target and identifies the site to which the address tag is delivered; analyzing the probe/address tag conjugate bound to the one or more biological targets, the analysis comprising;
(1) determining abundance, expression, and/or activity of each of the one or more biological targets by assessing the amount of the probe/address tag conjugate bound to the biological target; and
(2) determining the identities of the identity tag and the address tag of the probe/address tag conjugate; anddetermining a spatial pattern of abundance, expression, and/or activity of the one or more biological targets across the multiple sites in the sample based on the analysis, wherein the method does not depend on an imaging technique for determining spatial information of the one or more biological targets in the sample.
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4. A method of determining a spatial pattern of abundance, expression, and/or activity of one or more biological targets across multiple sites in a sample, comprising:
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delivering a probe for each of one or more biological targets to multiple sites in a sample, wherein each probe comprises;
(1) a target-binding moiety capable of binding to the probe'"'"'s corresponding biological target; and
(2) an identity tag having a sequence that identifies the probe'"'"'s corresponding biological target or target-binding moiety;allowing each probe to bind to its corresponding biological target in the sample; delivering an address tag to each of the multiple sites in the sample, wherein the address tag is to be coupled to the probe bound to the biological target and identifies the site to which the address tag is delivered; analyzing the probe/address tag conjugate bound to the one or more biological targets by sequencing, wherein the amount of a sequencing product indicates abundance, expression, and/or activity of each of the one or more biological targets, the sequencing product comprising all or a portion of the address tag sequence and all or a portion of the identity tag sequence; and determining a spatial pattern of abundance, expression, and/or activity of the one or more biological targets across the multiple sites in the sample based on the analysis.
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12. A method of determining a spatial pattern of abundance, expression, and/or activity of a target protein across multiple sites in a sample, comprising:
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delivering a probe for a target protein to multiple sites in a sample, wherein the probe comprises;
(1) a target-binding moiety capable of binding to the target protein;
(2) a first address tag that identifies each of the multiple sites to which the probe is delivered; and
(3) an identity tag that identifies the target protein or the target-binding moiety;allowing the probe to bind to the target protein in the sample; analyzing the probe bound to the target protein, the analysis comprising;
(1) determining abundance, expression, and/or activity of the target protein by assessing the amount of the probe bound to the target protein; and
(2) determining the identities of the identity tag and the first address tag of the probe for the target protein; anddetermining a spatial pattern of abundance, expression, and/or activity of the target protein across the multiple sites in the sample based on the analysis. - View Dependent Claims (13, 16, 17, 18, 19, 20)
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14. A method of determining a spatial pattern of abundance, expression, and/or activity of a target protein across multiple sites in a sample, comprising:
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delivering a probe for a target protein to multiple sites in the sample, wherein the probe comprises;
(1) a target-binding moiety capable of binding to the target protein; and
(2) an identity tag that identifies the target protein or the protein-binding moiety;allowing the probe to bind to the target protein in the sample; delivering a first address tag to each of the multiple sites in the sample, wherein the first address tag is to be coupled to the probe bound to the target protein and identifies the site to which it is delivered; analyzing the probe/first address tag conjugate bound to the target protein, the analysis comprising;
(1) determining abundance, expression, and/or activity of the target protein by assessing the amount of the probe/first address tag conjugate bound to the target protein; and
(2) determining the identities of the identity tag and the first address tag of the probe/first address tag conjugate; anddetermining a spatial pattern of abundance, expression, and/or activity of the target protein across the multiple sites in the sample based on the analysis. - View Dependent Claims (15)
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52. A system for determining a spatial pattern of abundance, expression, and/or activity of one or more biological targets across multiple sites in a sample, comprising:
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a first module for delivering a probe for each of one or more biological targets to multiple sites in a sample, wherein each probe comprises;
(1) a target-binding moiety capable of binding to the probe'"'"'s corresponding biological target; and
(2) an identity tag that identifies the probe'"'"'s corresponding biological target or target-binding moiety;a second module for delivering an address tag to each of the multiple sites in the sample, wherein the address tag is to be coupled to the probe bound to the biological target and identifies the site to which the address tag is delivered; a third module for analyzing the probe/address tag conjugate bound to the one or more biological targets, the analysis comprising;
(1) determining abundance, expression, and/or activity of the one or more biological targets by assessing the amount of the probe/address tag conjugate bound to the biological target; and
(2) determining the identities of the identity tag and the address tag of the probe/address tag conjugate; anda fourth module for determining a spatial pattern of abundance, expression, and/or activity of the one or more biological targets across the multiple sites in the sample based on the analysis, wherein the system does not depend on an imaging technique for determining spatial information of the one or more biological targets in the sample. - View Dependent Claims (53, 54, 55, 56)
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57. A method of determining a spatial pattern of abundance, expression, and/or activity of one or more biological targets across multiple sites in a sample, comprising:
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delivering a probe for each of one or more biological targets to multiple sites in a sample, wherein each probe comprises a target-binding moiety capable of binding to the probe'"'"'s corresponding biological target; allowing each probe to bind to its corresponding biological target in the sample; delivering at least one adaptor to the multiple sites in the sample, wherein the at least one adaptor specifically binds to the probe and comprises an address tag that identifies each of the multiple sites to which the at least one adaptor is delivered, wherein the probe and/or the adaptor comprises an identity tag that identifies the probe'"'"'s and/or adaptor'"'"'s corresponding biological target or target-binding moiety; analyzing the at least one adaptor and the probe bound to the one or more biological targets, the analysis comprising;
(1) determining abundance, expression, and/or activity of each of the one or more biological targets by assessing the amount of at least one adaptor bound to the probe bound to the biological target; and
(2) determining the identities of the identity tag, and the address tag of the at least one adaptor; anddetermining a spatial pattern of abundance and/or activity of the one or more biological targets across the multiple sites in the sample based on the analysis, wherein the method does not depend on an imaging technique for determining spatial information of the one or more biological targets in the sample. - View Dependent Claims (61, 62, 63, 64)
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58. A method of determining a spatial pattern of abundance, expression, and/or activity of one or more biological targets across multiple sites in a sample, comprising:
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delivering a probe for each of one or more biological targets to multiple sites in a sample, wherein each probe comprises a target-binding moiety capable of binding to the probe'"'"'s corresponding biological target; allowing each probe to bind to its corresponding biological target in the sample; delivering at least one adaptor to the multiple sites in the sample, wherein the at least one adaptor specifically binds to the probe and comprises an address tag that identifies each of the multiple sites to which the at least one adaptor is delivered, wherein the probe and/or the adaptor comprises an identity tag that identifies the probe'"'"'s and/or adaptor'"'"'s corresponding biological target or target-binding moiety; analyzing the at least one adaptor and the probe bound to the one or more biological targets by sequencing, wherein the amount of a sequencing product indicates abundance, expression, and/or activity of each of the one or more biological targets, the sequencing product comprising all or a portion of the address tag sequence and all or a portion of the identity tag sequence; and determining a spatial pattern of abundance, expression, and/or activity of the one or more biological targets across the multiple sites in the sample based on the analysis, wherein the method does not depend on an imaging technique for determining spatial information of the one or more biological targets in the sample.
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59. A method of determining a spatial pattern of abundance, expression, and/or activity of one or more biological targets across multiple sites in a sample, comprising:
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delivering a probe for each of one or more biological targets to multiple sites in a sample, wherein each probe comprises a target-binding moiety capable of binding to the probe'"'"'s corresponding biological target; allowing each probe to bind to its corresponding biological target in the sample; delivering at least one adaptor to the multiple sites in the sample, wherein the at least one adaptor specifically binds to the probe, wherein the probe and/or the adaptor comprises an identity tag that identifies the probe'"'"'s and/or adaptor'"'"'s corresponding biological target or target-binding moiety; delivering an address tag to each of the multiple sites in the sample, wherein the address tag is to be coupled to the at least one adaptor bound to the probe bound to the biological target and identifies the site to which the address tag is delivered; analyzing the adaptor/address tag conjugate, the analysis comprising;
(1) determining abundance, expression, and/or activity of each of the one or more biological targets by assessing the amount of the adaptor/address tag conjugate bound to the probe bound to the biological target; and
(2) determining the identities of the identity tag, and the address tag of the adaptor/address tag conjugate; anddetermining a spatial pattern of abundance, expression, and/or activity of the one or more biological targets across the multiple sites in the sample based on the analysis, wherein the method does not depend on an imaging technique for determining spatial information of the one or more biological targets in the sample.
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60. A method of determining a spatial pattern of abundance, expression, and/or activity of one or more biological targets across multiple sites in a sample, comprising:
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delivering a probe for each of one or more biological targets to multiple sites in a sample, wherein each probe comprises a target-binding moiety capable of binding to the probe'"'"'s corresponding biological target; allowing each probe to bind to its corresponding biological target in the sample; delivering at least one adaptor to the multiple sites in the sample, wherein the at least one adaptor specifically binds to the probe, wherein the probe and/or the adaptor comprises an identity tag that identifies the probe'"'"'s and/or adaptor'"'"'s corresponding biological target or target-binding moiety; delivering an address tag to each of the multiple sites in the sample, wherein the address tag is to be coupled to the at least one adaptor bound to the probe bound to the biological target and identifies the site to which the address tag is delivered; analyzing the adaptor/address tag conjugate by sequencing, wherein the amount of a sequencing product indicates abundance, expression, and/or activity of each of the one or more biological targets, the sequencing product comprising all or a portion of the address tag sequence and all or a portion of the identity tag sequence; and determining a spatial pattern of abundance, expression, and/or activity of the one or more biological targets across the multiple sites in the sample based on the analysis.
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Specification