Assay devices and methods of use thereof
Inventors
Ririe, Kirk M. • Wernerehl, Aaron D • Pasko, Christopher P. • Laayoun, Ali • Vachon, Carole • Dupont-Filliard, Agnès • Mesta, Laurent • Hatch, Andrew C. • Huynh, Erik W. • Jones, David E.
Assignees
Biomerieux SA • Biofire Defense LLC • Biofire Diagnostics LLC
Publication Number
US-11691152-B2
Publication Date
2023-07-04
Expiration Date
2038-05-23
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Abstract
Systems, methods, and apparatuses are provided for self-contained nucleic acid preparation, amplification, and analysis.
Core Innovation
The invention provides systems, methods, and apparatuses for self-contained nucleic acid preparation, amplification, and analysis, utilizing a closed reaction vessel—such as flexible pouches or cards—designed to house all necessary reagents and components for the process. These self-contained reaction vessels incorporate zones for sample preparation, nucleic acid extraction, first-stage amplification (such as PCR), and second-stage amplification, each fluidly connected within the vessel. The vessel may be fabricated from flexible and/or rigid film layers and can be provided with liquid or dried reagents in blisters at the time of manufacture.
The problem addressed is that existing diagnostic methods for infectious diseases, specifically PCR-based detection, face issues such as the need for multiple, separate assays for different pathogens, low pathogen concentration in samples, inadequate sample volumes for comprehensive testing, delays due to manual handling and multi-step protocols, and contamination risk during open handling. Multiplex PCR methods, though helpful, are limited by challenging analysis and robustness for high-level multiplexing. There is a need for a rapid, automated, cost-effective, and user-friendly system to process diverse sample types with minimized user input and contamination risk.
The core innovation includes a reaction vessel with one or more multifunction chambers for integrated sample lysis, nucleic acid binding (using magnetic or other nucleic acid-binding particles), washing, and amplification. First-stage amplification (singleplex or multiplex) is followed by fluent transfer of a reaction aliquot to a second-stage reaction zone containing multiple discrete wells, each with immobilized primer sets for further amplification and simultaneous thermal cycling. Reagent blisters, openable seals (such as burstable or tacked seals), and accessories like sample collection swabs are included in the vessel to automate the workflow from sample input to answer, compatible with associated instrumentation for pneumatic or mechanical fluid movement, heating, cooling, and optical detection.
Claims Coverage
The independent claim covers five main inventive features that define the core of the invention.
Self-contained nucleic acid amplification vessel with multifunction chamber
A container comprising a first and second flexible barrier film layer that defines a multifunction chamber between them, where the chamber is provided with magnetic particles and reagents for sample preparation, nucleic acid recovery, and a first-stage nucleic acid amplification reaction. The magnetic particles and reagents are supplied in one or more fluid-filled reagent blisters that are filled at the time of manufacture, and the magnetic particles may be housed in or fluidly connected to the multifunction chamber.
Second-stage reaction zone with plurality of reaction chambers for further amplification
A second-stage reaction zone is disposed between the film layers and is fluidly connected to the multifunction chamber. This zone consists of multiple second-stage reaction chambers, each comprising a pair of primers for further amplification of the sample. All second-stage chambers are configured for contemporaneous thermal cycling, and are different from the chamber used for first-stage nucleic acid amplification.
Flexible barrier film construction with specified water vapor transmission rate
The first and second layers of the container are flexible barrier films with a specified water vapor transmission rate (WVTR) in the range of about 0.05 g/m2/24 hrs to about 2 g/m2/24 hrs, ensuring suitable barrier properties for biological sample and assay stability.
Inclusion of sample receiving chamber and swab-based sample input option
The container may further comprise a sample receiving chamber in fluid communication with the multifunction chamber. The chamber may include a sample collection swab with an elongate shaft, and both the swab and the sample receiving chamber are fabricated from chemically compatible materials that can be at least partially fused with a heat seal.
Use of openable seals for controlled reagent delivery
An openable seal (such as a burstable seal or a peelable, tacked together film seal) exists between the fluid-filled reagent blisters and the multifunction chamber, enabling controlled release of reagents into the chamber at the appropriate stage during the assay process.
The independent claim defines a closed-system container for nucleic acid sample processing and amplification, emphasizing the integration of reagent blisters, sample-to-answer automation, consecutive amplification stages, specialized film construction, and user-friendly, sealed sample input.
Stated Advantages
The invention enables rapid sample-to-answer processing by integrating sample preparation, nucleic acid amplification, and analysis in a single, self-contained vessel.
Automation and sealed workflow reduce user input and minimize contamination risk during processing of clinical or environmental samples.
Flexible and/or rigid container designs allow for cost-effective manufacturing by highly automated methods and facilitate simple packaging.
On-board storage of reagents (liquid or dried) at the time of manufacture simplifies operation by end users and enhances ease of use.
The system supports a wide range of sample types and sample collection methods with minimal manual manipulation.
Documented Applications
Diagnosis of infectious diseases through multiplexed nucleic acid detection from diverse clinical sample types such as blood, nasopharyngeal swabs, sputum, stool, and more.
Environmental and veterinary pathogen nucleic acid detection, including testing of environmental samples like soil, water, air monitoring filters, and vectors such as mosquitos or ticks.
Use in point-of-care, laboratory, and automated diagnostic instruments for analyzing pathogen or organism nucleic acids with minimal manual input.
High-throughput screening for multiple pathogens or targets in a single reaction vessel using multiplex and nested PCR workflows.
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