Blood sample assay method
Inventors
Faaren, Arne Ludvig • Frantzen, Frank • Nordhei, Arne Kristian • Sundrehagen, Erling • Ørning, Lars
Assignees
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Publication Number
US-11814669-B2
Publication Date
2023-11-14
Expiration Date
Abstract
The invention provides an enzymatic method for measuring the concentration of one or more analytes in the plasma portion of a blood derived sample, containing a first and a second component, where said second component interferes with the measurement of said first component. The method includes: i) diluting the sample with a reagent mixture; ii) substantially removing blood cells; iii) using a reagent which serves to temporarily prevent reaction of the second component, to generate a blocked second component; iv) causing the selective reaction of a constituent of each analyte to directly or indirectly generate detectable reaction products, where one of the analytes is the first component; v) monitoring the detectable reaction product or products; vi) relating an amount of the detectable product or products and/or a rate of formation of the detectable product or products to the concentration of each analyte, where the concentration of at least the first component is related to a corresponding detectable reaction product by means of estimating an un-measurable (fictive) endpoint. Step iii) may be carried out at any stage up to and including step iv) but before steps v) or vi). The reagent of step iii) may be applied to the sample separately or may be included in a reagent mixture during steps i) or iv). A corresponding kit is also provided.
Core Innovation
The invention relates to an enzymatic point-of-care blood assay for plasma lipid/analyte measurement in which an interfering second component is temporarily blocked and cells are removed to obtain a cell-free portion. Because the targeted reaction lacks a measurable endpoint due to the temporary blocking and unblocking or blocking and unreactive behavior, the assay estimates a fictive endpoint from partial reaction progress based on a reaction progress curve and curve-fitting algorithms, and uses the estimated fictive endpoint to determine a detectable reaction product signal for the targeted assay readout.
The invention provides an HDL assay correction approach that uses a total cholesterol (CH) value and a measured HDL value for the sample. A non-HDL value is calculated by subtracting the measured HDL value from the CH value, and a corrected HDL value is calculated using a function relating the non-HDL value and an increase in HDL, wherein the corrected HDL value is calculated by subtracting the increase in HDL from the measured HDL value.
The described workflow and kit integrate sample handling and assay components for converting analytes to detectable products, including dilution reagent, a cell separation unit for cell removal such as filtration, and a temporary blocking reagent that selectively blocks an interfering second component while an enzymatic conversion proceeds. The documented approach includes using fitted reaction progress data to estimate an endpoint absorbance or fictive, unmeasurable, unreachable endpoint and uses correction and calibration concepts to relate non-HDL to an increase in HDL, enabling corrected HDL determination from the measured values.
Claims Coverage
Independent claim clm-00001 covers a method for correcting an HDL assay by combining a CH value, a measured HDL value, a non-HDL calculation, and a correction that subtracts an HDL increase derived from a function relating the non-HDL value and the increase in HDL. Dependent claims refine how the measured HDL value is obtained and how the correction function is defined and applied.
Correcting HDL assay using CH and measured HDL values
obtaining a total cholesterol (CH) value for a sample; obtaining a measured high density lipoprotein (HDL) value for the sample; calculating a non-HDL value by subtracting the measured HDL value from the CH value; calculating a corrected HDL value using a function relating the non-HDL value and an increase in HDL, wherein said corrected HDL value is calculated by subtracting said increase in HDL from said measured HDL value.
Estimating an unmeasurable or fictive HDL endpoint
The measured HDL value is obtained by estimating an unmeasurable or fictive endpoint of an HDL assay performed on the sample.
Using a calibration sample with known HDL and non-HDL to determine HDL increase
providing a calibration sample with known HDL and non-HDL; measuring HDL in that calibration sample; calculating the increase in measured HDL by subtracting the known HDL amount from the measured HDL value.
Employing an exponential correlation for the HDL correction function
a function used in the method describes an exponential correlation between an increase in HDL and a non-HDL value.
Deriving HDL from end-point absorbance estimated from fitted reaction progress
determining a measured HDL value by estimating an end-point absorbance from a fitted curve to part of a reaction progress curve.
Using spectrophotometry to measure HDL value
measuring the HDL value using spectrophotometry.
Overall, the claim set supports correcting a measured HDL value by deriving non-HDL from CH minus measured HDL and then applying a function that relates non-HDL to an increase in HDL; the correction is refined by defining the measured HDL via fictive or unmeasurable endpoint estimation from reaction progress fitting, including end-point absorbance estimation, and by defining the correction function using an exponential correlation and calibration concepts, with spectrophotometry as a measurement modality in at least one refinement.
Stated Advantages
Enables corrected HDL determination using CH and measured HDL values with a non-HDL-based correction function.
Allows HDL assay operation where the targeted reaction lacks a measurable endpoint by estimating a fictive or unmeasurable endpoint from partial reaction progress.
Improves assay time and supports minimal sample volume.
Reduces assay-time dependence on reagent storage.
Minimizes cell lysis via ionic-strength control.
Enables whole blood filtration to obtain a cell-free filtrate.
Enables multi-analyte lipid panels and may correct HDL estimation for parallel non-HDL conversion via post-analysis.
Documented Applications
Point-of-care blood assay use for plasma lipid/analyte measurement, including HDL and non-HDL/cholesterol-related lipid panel measurements.
Correcting a high density lipoprotein (HDL) assay by using total cholesterol (CH) and a measured HDL value to calculate and apply a corrected HDL value.
Use with enzymatic conversion workflows that incorporate temporary blocking of an interfering second component and cell separation or removal to obtain a cell-free portion, including whole blood filtration to obtain cell-free filtrate.
Use for determining HDL measurement from reaction progress curve fitting, including estimating end-point absorbance from part of a reaction progress curve, when a measurable endpoint is not available.
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