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Publication Number
US-9618520-B2
Publication Date
2017-04-11
Expiration Date
Abstract
Microarray compositions suitable for analysis by one or several spectrographic methods are disclosed. In an embodiment, a microarray composition includes a three-dimensional solid support and a plurality of reactive microbeads positioned on the solid support in spatially distinct and addressable locations.
Core Innovation
The invention provides a composite microarray that includes a three-dimensional solid support having a first analytical site and a first reactive site distinct from the first analytical site. The first reactive site includes a first bead associated with at least one type of active agent configured to bind a compound, and the first analytical site and the first reactive site are fluidically connected so that at least one analyte migrates from the first bead to the first analytical site.
The at least one analyte includes at least one member selected from the group consisting of the active agent, a fragment of the active agent, a chemically modified form of the active agent, an enzymatically modified form of the active agent, and a compound bound to the active agent. Dimensions of the first analytical site do not exceed 1 mm, and the geometry supports distinctness between the analytical site and the reactive site while enabling analyte migration and localization for readout.
The disclosure also provides high-throughput single-cell whole-cell mass spectrometry and mass-spectrometric imaging enabled by densely packed microwell or fiber-optic microwell plates. It describes reproducible MALDI TOF MS results and data-processing approaches for comparing spectra, including difference mass spectrometry using a Sample Spectrum minus a Reference Spectrum, with least squares subtraction.
A related method analyzes a sample by providing a suspension of reactive sites, each reactive site including a bead with at least one active agent configured to bind a compound. The contacted suspension is distributed into an array of microwells of a microwell plate to form a composite microarray, and after allowing reactive sites to react with the sample and removing an unreacted portion, captured analytes migrate from the beads to analytical sites that are distinct from the reactive sites.
Claims Coverage
The partial content includes two independent claims: a composite microarray claim and a method of analyzing a sample claim. Across the independent claims, the core inventive features include distinct analytical and reactive sites on a solid support or microwell plate, fluidic connectivity to enable analyte migration, bead-associated active agents configured to bind a compound, analytical sites with dimensions not exceeding 1 mm, and spectrographic analysis of localized analytes.
Distinct analytical and reactive sites on a composite microarray
A composite microarray comprising a solid support including a first analytical site and a first reactive site associated with the solid support, the first reactive site comprising a first bead and at least one type of active agent configured to bind a compound, wherein the dimensions of the first analytical site do not exceed 1 mm and the first analytical site is distinct from the first reactive site.
Fluidic connection enabling analyte migration to the analytical site
The first reactive site is fluidically connected to the first analytical site so as to enable at least one analyte to migrate from the first bead to the first analytical site, where the at least one analyte is at least one member selected from the group consisting of the active agent, a fragment of the active agent, a chemically modified form of the active agent, an enzymatically modified form of the active agent, and a compound bound to the active agent.
Forming a composite microarray from reactive sites in microwells
A method of analyzing a sample comprising providing a suspension of reactive sites, contacting the sample with the suspension, and distributing the contacted suspension into an array of microwells of a microwell plate whereby a composite microarray is formed, wherein each reactive site comprises a bead and at least one type of active agent configured to bind a compound.
Migration of captured analytes to distinct analytical sites for spectrographic analysis
Allowing the reactive sites to react with the sample, removing an unreacted portion of the sample from the composite microarray, effectuating migration of a captured analyte from the beads of the reactive sites to analytical sites that are distinct from the reactive sites, wherein dimensions of each analytical site do not exceed 1 mm, and analyzing the analytical sites by a spectrographic method.
The independent claims cover a composite microarray with distinct analytical and reactive sites on a solid support, where bead-associated active agents bind compounds and analytes migrate via fluidic connection from reactive sites to distinct analytical sites. A method claim further specifies forming the composite microarray by distributing reactive-site-containing suspensions into an array of microwells, removing unreacted sample, enabling migration to dimension-limited analytical sites, and analyzing those sites by a spectrographic method.
Stated Advantages
High-throughput single-cell whole-cell mass spectrometry and mass-spectrometric imaging.
Reproducible MALDI TOF MS results, including detection of even low-abundance peaks.
Supports quantitative mass-spectrometric readout.
Documented Applications
Cell-level and rare-cell detection contexts, including drug- and apoptosis-related responses.
Mass-spectrometric imaging.
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