Spatially encoded biological assays
First Claim
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1. A method to determine the abundance or activity or both of a biological target in a sample, comprising the following steps:
- delivering encoded probes for a biological target to multiple sites in a sample affixed to a support, wherein each encoded probe comprises a probe region capable of specific binding to the biological target and a coding tag comprising an oligonucleotide that identifies the location of the site to which the encoded probe is delivered;
allowing the encoded probes to interact with the biological target;
separating encoded probes specifically bound to the biological target from encoded probes not specifically bound to the biological target;
pooling encoded probes specifically bound to the biological target;
determining all or a portion of the coding tag oligonucleotide sequence of the encoded probe in the pool by performing a polynucleotide sequencing reaction of the encoded probe, thereby identifying the location of the site to which the encoded probe is delivered; and
associating the abundance or activity or both of the biological target to the locations of the multiple sites in the sample.
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Abstract
The present invention provides assays and assay systems for use in spatially encoded biological assays. The invention provides an assay system comprising an assay capable of high levels of multiplexing where reagents are provided to a biological sample in defined spatial patterns; instrumentation capable of controlled delivery of reagents according to the spatial patterns; and a decoding scheme providing a readout that is digital in nature.
250 Citations
46 Claims
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1. A method to determine the abundance or activity or both of a biological target in a sample, comprising the following steps:
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delivering encoded probes for a biological target to multiple sites in a sample affixed to a support, wherein each encoded probe comprises a probe region capable of specific binding to the biological target and a coding tag comprising an oligonucleotide that identifies the location of the site to which the encoded probe is delivered; allowing the encoded probes to interact with the biological target; separating encoded probes specifically bound to the biological target from encoded probes not specifically bound to the biological target; pooling encoded probes specifically bound to the biological target; determining all or a portion of the coding tag oligonucleotide sequence of the encoded probe in the pool by performing a polynucleotide sequencing reaction of the encoded probe, thereby identifying the location of the site to which the encoded probe is delivered; and associating the abundance or activity or both of the biological target to the locations of the multiple sites in the sample. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14)
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15. A method to determine the abundance or activity or both of a nucleic acid target in a sample, comprising the following steps:
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delivering an oligonucleotide probe capable of specific hybridization with a nucleic acid target to multiple sites in a sample affixed to a support; allowing the oligonucleotide probe to hybridize with the nucleic acid target; washing unhybridized oligonucleotide probe from the sample; delivering one or more encoding agents to the multiple sites in the sample, wherein the one or more encoding agents delivered to each site comprise an oligonucleotide that identifies the location of the site in the sample; coupling the one or more encoding agents and the oligonucleotide probe delivered to each site to form an encoded probe specifically hybridized with the nucleic acid target at the site; pooling encoded probes specifically hybridized with the nucleic acid target; determining all or a portion of the one or more encoding agent oligonucleotide sequences of the encoded probe in the pool by performing a polynucleotide sequencing reaction of the encoded probe, thereby identifying the location of the site to which the one or more encoding agents are delivered; and associating the abundance or activity or both of nucleic acid target to the locations of the multiple sites in the sample. - View Dependent Claims (16, 17, 18, 19, 20, 21, 22)
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23. A method of determining the abundance or activity or both of a biological target in a sample, comprising:
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delivering a probe capable of specific binding to a biological target to multiple sites in a sample affixed to a support; allowing the probe to interact with the biological target at each of the multiple sites in the sample; delivering an encoding agent to each of the multiple sites in the sample, wherein the encoding agent delivered to each site comprises an oligonucleotide that identifies the location of the site in the sample; coupling the encoding agent to the probe delivered to each of the multiple sites to form an encoded probe at each of the multiple sites; pooling encoded probes specifically bound to the biological target; determining all or a portion of the encoding agent oligonucleotide sequence of the encoded probe in the pool by performing a polynucleotide sequencing reaction of the encoded probe, thereby identifying the location of the site to which the encoding agent is delivered; and associating the abundance or activity or both of the biological target at each of the multiple sites to the locations of the multiple sites in the sample. - View Dependent Claims (24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46)
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Specification