Fluorescence Polarization Immunoassay Calibration Against Autofluorescence
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Solution Overview
Problem
The influence of autofluorescent materials in samples interferes with the accurate measurement of target substance concentration in fluorescence polarization immunoassays, making it difficult to obtain the intrinsic degree of polarization in competitive FPIA.
Innovation Solution
A method and device that corrects the degree of polarization by using reference samples with known target substance concentrations, allowing for the generation of calibration curves that account for autofluorescence, using a fluorescence polarization immunoassay device with a polarization adjustment element and controller to measure and correct fluorescence intensities.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If fluorescence polarization measurement is performed in competitive FPIA, then target substance concentration can be quantified, but measurement accuracy deteriorates due to autofluorescence interference
Solution Approach 1:
The patent extracts the autofluorescence component from the total fluorescence signal by measuring a blank sample (without tracer) separately. This isolated autofluorescence value is then subtracted from the total polarization signal to obtain the corrected tracer-specific polarization value, thereby eliminating the harmful interference effect.
Solution Approach 2:
The patent introduces a blank sample (first reference sample) as an intermediary measurement. This blank sample contains all components except the fluorescent tracer, serving as a mediator to quantify and subsequently remove the autofluorescence contribution from the total signal in the test samples.
2Productivity
If conventional FPIA calibration curves are used, then quantification can be performed, but measurement reliability deteriorates due to uncorrected autofluorescence
Solution Approach 1:
The patent performs preliminary correction of the calibration curve by measuring and subtracting autofluorescence from all calibration standards using the same blank sample correction method. This preliminary action ensures that the calibration curve itself is free from autofluorescence interference, making subsequent measurements reliable.
Solution Approach 2:
The patent implements a feedback mechanism where the blank sample measurement provides correction information that is fed back into both the calibration curve generation and the test sample measurement process. This feedback loop ensures consistent correction across all measurements, improving overall reliability.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The method and device effectively reduce measurement errors caused by autofluorescence, enabling accurate quantification of target substance concentrations by generating corrected calibration curves.
Implementation Method 1
a fluorescently labeled substance, obtained by labeling the target substance with a fluorochrome
Implementation Method 2
fluorescence polarization measurement
Data Source
AI summary
A degree of polarization of second reference samples produced by adding an antibody, a tracer, and a target substance having mutually different concentrations to a first reference sample not including the target substance is corrected with a degree of polarization of the first reference sample to generate a first calibration curve. A degree of polarization of a second sample to be measured produced by adding the antibody and the tracer to a first sample to be measured in amounts equal to those added to the second reference samples is corrected with a degree of polarization of the first sample to be measured to obtain the concentration of the target substance in the first calibration curve.