Apparatus, systems, and methods for quantifying infectious agents
Inventors
RAJAN, Nitin K. • KNOPFMACHER, Oren S. • Herget, Meike
Assignees
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Publication Number
US-12379373-B2
Publication Date
2025-08-05
Expiration Date
Abstract
Various methods, devices, and systems for determining the concentration of infectious agent in a target sample are disclosed herein. In one embodiment, a method for determining the concentration of an infectious agent of an unknown strain can include diluting aliquots of a target sample comprising the infectious agent by different dilution factors to yield diluted samples. The method can also include determining the time it takes a solution characteristic of each of the diluted samples to undertake a predetermined threshold change. The method can also include determining the concentration of the infectious agent of the unknown strain by taking into account the different dilution factors, the monitored times, and certain curve fitting parameters calculated from predetermined calibration curves generated for infectious agents of different known strains.
Core Innovation
The invention provides methods and systems for in vitro quantification of a bacteria of an unknown strain in a target sample by diluting the target sample with two dilution factors and measuring oxidation reduction potential using corresponding ORP sensors. The ORP is monitored in the absence of any added reporter molecules while determining first and second times-to-detection that represent the time for the ORP of each diluted sample to undertake a predetermined threshold change.
A calibration framework is used that computes an average calibration curve slope and an average calibration curve y-intercept from multiple calibration curves representing growth behavior of one or more bacteria of different known strains. Corrected calibration parameters are then calculated, including a corrected calibration curve slope using at least the first and second times-to-detection and the two dilution factors, and a corrected calibration curve y-intercept using at least the average y-intercept, the corrected slope, and the average slope.
Finally, the concentration of the bacteria of the unknown strain in the target sample is determined using at least the corrected calibration parameters and either the first time-to-detection with the first dilution factor or the second time-to-detection with the second dilution factor. In examples, solution characteristic measurement is described for ORP using redox-active sensor materials and for pH using threshold pH changes and functionalization layers, including combined ORP/pH sensor architectures.
Claims Coverage
The document contains one independent claim. The inventive features focus on dual dilution with two ORP measurements without added reporter molecules, time-to-detection based on a predetermined ORP threshold change, calibration using multiple known-strain calibration curves to compute average and corrected calibration parameters, and calculating the unknown bacterial concentration from corrected calibration parameters and the selected time/dilution inputs.
Dual dilution ORP measurement without reporter molecules
Diluting a first aliquot of the target sample by a first dilution factor to yield a first diluted sample; diluting a second aliquot by a second dilution factor to yield a second diluted sample; exposing the first diluted sample to a first oxidation reduction potential sensor and the second diluted sample to a second oxidation reduction potential sensor, each comprising a redox-active material, wherein the ORP is monitored in the absence of any added reporter molecules in any of the first diluted sample or the second diluted sample.
Time-to-detection from a predetermined ORP threshold change
Determining a first time-to-detection representing the time it takes the ORP of the first diluted sample to undertake a predetermined threshold change; determining a second time-to-detection representing the time it takes the ORP of the second diluted sample to undertake the predetermined threshold change.
Average and corrected calibration from multiple known-strain calibration curves
Calculating an average calibration curve slope and an average calibration curve y-intercept from equation parameters obtained from multiple calibration curves representing growth behavior of one or more bacteria of different known strains; calculating a corrected calibration curve slope using at least the first and second times-to-detection and the two dilution factors; calculating a corrected calibration curve y-intercept using at least the average y-intercept, the corrected slope, and the average slope.
Concentration determination using corrected calibration parameters
Determining the concentration of the bacteria of the unknown strain in the target sample using at least the corrected calibration parameters and either the first time-to-detection and the first dilution factor or the second time-to-detection and the second dilution factor.
Claim coverage centers on using two dilution factors and two ORP sensors without added reporter molecules to obtain first and second times-to-detection, then applying calibration derived from multiple known strains to compute average and corrected calibration parameters, and finally determining the unknown concentration using the corrected calibration parameters together with the corresponding time and dilution inputs.
Stated Advantages
Enables determining a concentration of a bacteria of an unknown strain in a target sample using ORP monitored without any added reporter molecules.
Uses calibration curves representing growth behavior of one or more bacteria of different known strains to compute corrected calibration parameters for concentration determination.
Documented Applications
In vitro quantification of a bacteria of an unknown strain in a target sample by dual dilution and ORP-based time-to-detection with calibration using known strains.
Measurement of a solution characteristic via ORP sensors including ORP monitoring in the absence of added reporter molecules.
Measurement of a solution characteristic via pH with threshold pH changes and functionalization layers, including combined ORP/pH sensor architectures [procedural detail omitted for safety].
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