High density self-contained biological analysis
Inventors
Ririe, Kirk M. • Poritz, Mark Aaron • Rasmussen, Randy P.
Assignees
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Publication Number
US-11866774-B2
Publication Date
2024-01-09
Expiration Date
Abstract
Devices, containers, and methods are provided for performing biological analysis in a closed environment. Illustrative biological analyses include high density nucleic acid amplification and detection and immuno-PCR.
Core Innovation
The invention describes nucleic acid extraction and multiplex PCR carried out in a self-contained system that includes one or more ports, an injector port for introducing the sample, a nucleic acid preparation zone for purifying nucleic acids, and at least one amplification zone configured for amplification of the plurality of nucleic acids that may be in the sample. The sample is introduced via the injector port and the injector port is closed subsequent to introducing the sample. Nucleic acids prepared in the nucleic acid preparation zone are moved into the amplification zone.
The invention further provides multiplex PCR by mixing the nucleic acids in the sample with PCR reaction components including a plurality of primer pairs configured for amplifying the targets. Thermal cycling is performed in the amplification zone to create an amplification mixture, and mixing continues during thermal cycling. Detection is then performed to determine which of the plurality of nucleic acids are present in the amplification mixture.
The disclosed system architecture additionally specifies container arrangements that include a cell lysis zone connected to the nucleic acid preparation zone, and amplification-zone configurations that support nested two-stage amplification within first-stage and second-stage amplification zones. The document also describes minimizing cross-contamination in high-density reaction wells/chambers using barrier layers. Immuno-PCR workflows are described in which antibody-DNA conjugates form complexes and PCR readout is performed within the closed pouch, including combined PCR and iPCR workflows.
Claims Coverage
The independent claim recites a method for performing nucleic acid extraction and multiplex PCR in a self-contained system, involving inventive features covering closed-port sample introduction, nucleic-acid purification, multiplex primer-based amplification, ongoing mixing during thermal cycling, and detection of which targets are present. No additional independent claims are explicitly provided in the partial content beyond clm-00001.
Self-contained extraction and multiplex PCR with injector port, nucleic acid preparation zone, and amplification zone
A method comprising providing the self-contained system having, in fluid communication, one or more ports including an injector port, a nucleic acid preparation zone configured for purifying nucleic acids, and at least one amplification zone configured for amplification of the plurality of nucleic acids that may be in the sample.
Closed injector port after sample introduction
Introducing the sample into the self-contained system via the injector port, and closing the injector port subsequent to introducing the sample into the self-contained system.
Purify and move nucleic acids into amplification zone
Preparing the nucleic acids in the nucleic acid preparation zone, and moving the nucleic acids into the amplification zone.
Multiplex primer-pair PCR with targets
Mixing the nucleic acids in the sample with PCR reaction components including a plurality of primer pairs configured for amplifying the targets.
Thermal cycling with mixing continuing during thermal cycling
Thermal cycling the nucleic acids in the amplification zone to create an amplification mixture, wherein mixing continues during thermal cycling.
Detect which nucleic acids are present
Detecting which of the plurality of nucleic acids are present in the amplification mixture.
Overall, the claim coverage centers on executing nucleic acid extraction and multiplex PCR in a self-contained system with closed-port handling, nucleic-acid purification followed by transfer to an amplification zone, multiplex primer-pair amplification with ongoing mixing during thermal cycling, and detecting which targets are present in the amplification mixture.
Stated Advantages
Not explicitly described in patent.
Documented Applications
diagnostics
antibiotic-resistance profiling
forensic SNP typing
Process controls for the nucleic-acid preparation and amplification workflows
immuno-PCR (iPCR) workflows combining ELISA-to-PCR readout within the closed pouch
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