Methods and systems for extending dynamic range in assays for the detection of molecules or particles
Inventors
Rissin, David M. • Fournier, David • Duffy, David C. • Walt, David R. • Kan, Cheuk W
Assignees
Interested in licensing this patent?
MTEC can help explore whether this patent might be available for licensing for your application.
Publication Number
US-12019072-B2
Publication Date
2024-06-25
Expiration Date
Abstract
Described herein are systems and methods for extending the dynamic range of assay methods and systems used for determining the concentration of analyte molecules or particles in a fluid sample. In some embodiments, a method comprises spatially segregating a plurality of analyte molecules in a fluid sample into a plurality of locations. At least a portion of the locations may be addressed to determine the percentage of said locations containing at least one analyte molecule. Based at least in part on the percentage, a measure of the concentration of analyte molecules in the fluid sample may be determined using an analog, intensity-based detection/analysis method/system and/or a digital detection/analysis method/system. In some cases, the assay may comprise the use of a plurality of capture objects.
Core Innovation
The invention relates to a method for determining a measure of the concentration of analyte molecules or particles in a fluid sample by spatially associating analyte with capture objects and then performing detection based on the fraction of capture objects that become positive after addressing. Capture objects each include a binding surface having affinity for at least one type of analyte molecule or particle, and at least some capture objects become associated with the analyte after exposure to a solution containing or suspected of containing the analyte. Binding ligands are associated with analyte associated with capture objects, and the associated binding ligands are exposed to a precursor labeling agent converted into a labeling agent that becomes immobilized at or near the capture object.
After converting and immobilizing the labeling agent, capture objects are addressed, and detecting for each capture object addressed determines the presence or absence of immobilized labeling agent. The measure of analyte concentration is determined based at least in part on determining a measure indicative of the number or fraction of the capture objects subjected to the addressing step that are associated with immobilized labeling agent. The document further incorporates statistical concentration determination, including Poisson distribution analysis, for concentration determination from the number or fraction of capture objects determined to be associated with immobilized labeling agent.
The described approach also supports regimes that combine digital detection based on the percentage of active locations/beads with analog intensity-based detection using signal intensity and calibration/linked analysis, with thresholding between regimes. The document further discusses calibration and correction concepts, including handling background signal and controlling for factors such as substrate depletion and photobleaching, and includes examples for PSA measurement in serum showing a working range and low LOD across concentrations.
Claims Coverage
One independent claim is described. It covers a workflow for inferring analyte concentration from the fraction of addressed capture objects that have immobilized labeling agent, with statistical concentration determination such as Poisson distribution analysis in dependent claims.
Analyte capture with binding surfaces and associated binding ligands
Exposing capture objects, each including a binding surface having affinity for at least one type of analyte molecule or particle, to a solution containing or suspected of containing the at least one type of analyte molecule or particle, and associating at least one binding ligand with at least some analyte associated with a capture object.
Precursor labeling agent conversion and immobilization at or near capture objects
Exposing the at least one associated binding ligand to a precursor labeling agent such that the precursor labeling agent is converted to a labeling agent that becomes immobilized at or near the capture object with which the binding ligand is associated.
Addressing capture objects and detecting presence or absence of immobilized labeling agent
Addressing at least some of the capture objects after exposure to the precursor labeling agent, and detecting for each capture object addressed the presence or absence of immobilized labeling agent.
Concentration determination from the number or fraction of positive addressed capture objects
Determining a measure of the concentration of the analyte in the fluid sample based at least in part on a measure indicative of the number or fraction of the capture objects subjected to the addressing step determined to be associated with immobilized labeling agent.
The independent claim covers determining analyte concentration by converting a precursor labeling agent into immobilized labeling agent at or near analyte-associated capture objects, addressing the capture objects, detecting labeling-agent presence or absence, and calculating concentration from the number or fraction of positive addressed capture objects.
Stated Advantages
Enables determining a measure of analyte concentration over an extended dynamic range by using the number or fraction of addressed capture objects associated with immobilized labeling agent.
Supports concentration determination including Poisson distribution analysis based on the number or fraction of capture objects determined to be associated with immobilized labeling agent.
Reports PSA measurement in serum with a working range and demonstrated low LOD.
Documented Applications
PSA measurement in serum using the described method.
Interested in licensing this patent?