Two-Photon Fluorescence Clinical Assay Matrix Interference
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Solution Overview
Problem
Current clinical chemistry assays face challenges with high costs due to expensive disposable cuvettes and sensitivity issues related to sample matrix interferences and assay volumes, particularly in photometric detection methods, which are not suitable for point-of-care or distributed in vitro diagnostic (IVD) applications.
Innovation Solution
The use of two-photon excited fluorescence (TPE) technology for clinical chemistry analyte quantification, allowing separation-free assays in microvolumes with low-cost disposable cuvettes and reduced assay volumes, while being tolerant to matrix interferences, enabling both clinical chemistry and immunoassays with a single instrument.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If photometric detection methods are used for clinical chemistry assays, then the assays can be performed with standard equipment, but the cost increases due to expensive disposable cuvettes and sensitivity decreases due to matrix interferences
Solution Approach 1:
The patent changes the detection parameter from absorbance measurement to fluorescence measurement, and specifically from one-photon to two-photon excited fluorescence. This parameter change enables the use of microvolume samples and reduces matrix interference effects while maintaining assay sensitivity
Solution Approach 2:
The patent transitions from conventional photometric detection in a planar cuvette to three-dimensional two-photon excited fluorescence detection using focused laser excitation. This dimensional change allows excitation and detection to be confined to a small focal volume, reducing the impact of matrix interferences from surrounding sample regions
2Measurement precision
If photometric detection with disposable cuvettes is used, then assay precision can be maintained, but the cost increases significantly due to expensive cuvettes
Solution Approach 1:
The patent extracts the detection function from the cuvette by using two-photon excited fluorescence with focused laser excitation and emitted fluorescence detection. This allows the use of simple, inexpensive cuvettes or even well plates without compromising measurement precision, as the detection is based on fluorescence signal rather than optical path quality
Solution Approach 2:
The patent enables the use of cheap disposable cuvettes or well plates by replacing photometric detection with two-photon excited fluorescence detection. The inexpensive containers no longer need to meet strict optical quality requirements since the measurement is based on fluorescence emission rather than light absorption through the cuvette walls
3Ease of operation
If conventional photometric assays are performed, then standard assay volumes can be used, but the cost and complexity increase for point-of-care applications
Solution Approach 1:
The patent changes the detection method to two-photon excited fluorescence, which has different optical properties that enable sensitive detection in microvolumes. The non-linear excitation and emission characteristics allow for reduced sample volumes while maintaining detection sensitivity
Solution Approach 2:
The patent uses three-dimensional focused laser excitation and fluorescence detection to enable microvolume assays. The spatial confinement of excitation to a small focal volume and detection of emitted fluorescence allows for sensitive measurements in reduced sample volumes, making the assay suitable for point-of-care applications
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
TPE technology significantly reduces assay costs, improves accuracy by minimizing matrix interference effects, and allows for high sensitivity assays in smaller volumes, making it suitable for distributed IVD and point-of-care settings without the need for expensive cuvettes or complex liquid handling.
Implementation Method 1
two-photon excited fluorescence
Implementation Method 2
measuring two-photon exited fluorescence
Data Source
AI summary
The invention relates to an in vitro diagnostic method for quantification of a clinical chemistry analyte from a clinical sample wherein the clinical chemistry analyte undergoes a chemical reaction or reactions with a reagent or reagents in one or several steps, or in a reaction sequence, or catalyzes a chemical reaction, or reactions, or a reaction in a reaction sequence of a reagent or reagents, in one or several steps, in a reaction system. The reaction or reactions or reaction sequence result in a change of a measurable property of a compound or compounds of said reaction or reactions or reaction sequence. Characteristic for the method is that said chemical reaction or reactions or reaction sequence results in formation of a two-photon fluorescent compound, or a change in two-photon fluorescence properties of the reaction system comprising at least one two-photon fluorescent compound, and the analyte is quantified by exciting said two-photon fluorescent compound or compounds and measuring two-photon exited fluorescence, and relating said measured fluorescence to method standardization data based on measurements obtained from reference material of said analyte. The present invention also relates to use of a fluorometric device employing two-photon fluorescence excitation for quantification of a clinical chemistry analytes. The present invention further relates to a system for quantification of clinical chemistry analytes from samples containing the analyte. Characteristic for the system is that it comprises a fluorometric device employing two-photon excited fluorescence for quantifying one or several clinical chemistry analytes, and a data processing unit with software for dedicated data reduction for quantification of the analyte or analytes using said fluorometric device. The present invention further relates to a software product for the system.