Luminescent Labeled Particle Kit for High-Precision Biological Measurement
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Solution Overview
Problem
Current fluorescence detection methods, particularly the SPF method, face challenges in achieving high-precision measurements across a wide concentration range of measurement target substances due to insufficient fluorescence intensity and the influence of antibodies like anti-serum albumin antibodies present in biological samples, leading to nonspecific adsorption and reduced measurement reliability.
Innovation Solution
A kit and method utilizing labeled particles with a first binding substance and a first blocking agent, along with a substrate having a second binding substance and a second blocking agent, where the blocking agents are different, to enhance measurement precision by avoiding antibody interference and maintaining high quantum yield and luminance, especially using compounds represented by Formula (2) to suppress concentration quenching and achieve high fluorescence intensity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If the SPF method is used to improve detection sensitivity, then fluorescence intensity is enhanced, but measurement precision deteriorates due to nonspecific adsorption of antibodies in biological samples
Solution Approach 1:
The patent applies preliminary action by pre-coating the magnetic bead surface and substrate with blocking agents (such as BSA or casein) before introducing the biological sample. This preliminary blocking prevents antibodies and other proteins in the sample from nonspecifically adsorbing to the magnetic beads and substrate during the measurement process, thereby eliminating the harmful effect of nonsspecific adsorption while maintaining the high sensitivity of the SPF method
Solution Approach 2:
The patent uses blocking agents as intermediary substances that mediate between the magnetic bead surface/substrate and the biological sample components. These blocking agents occupy the binding sites on the magnetic beads and substrate, preventing direct nonsspecific interaction with antibodies in the sample, thus resolving the contradiction between maintaining surface availability for specific binding and preventing harmful nonsspecific adsorption
2Measurement precision
If particle concentration is reduced to improve detection sensitivity, then signal-to-noise ratio improves, but fluorescence intensity becomes insufficient
Solution Approach 1:
The patent applies parameter changes by optimizing the particle concentration to a specific low range (0.01-1×10^6 particles/mL) and compensating for the reduced fluorescence intensity through enhanced signal detection methods. The low particle concentration improves the signal-to-noise ratio by reducing background noise, while the SPF method's inherent sensitivity compensates for the lower total fluorescence signal, achieving both high detection sensitivity and sufficient signal intensity
Solution Approach 2:
The patent replaces conventional epi-fluorescence detection with surface plasmon fluorescence (SPF) detection, which uses surface plasmon resonance to enhance the fluorescence signal. This substitution of detection mechanism provides approximately 10-fold signal enhancement, allowing sufficient fluorescence intensity to be achieved even at very low particle concentrations, thereby resolving the contradiction between low particle concentration and sufficient fluorescence intensity
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
This approach enables high-precision measurement of measurement target substances across a wide concentration range from low to high concentrations, effectively overcoming the limitations of nonspecific adsorption and improving sensitivity and reliability.
Implementation Method 1
a luminescent labeled particle containing at least one kind of compound represented by Formula (2) and a particle
Implementation Method 2
The surface plasmon is generated in the metal layer by the irradiation with the excitation light, and the signal/noise ratio (S/N ratio) is improved by fluorescence enhancement, which is induced by the electric field enhancement effect caused by generation of the surface plasmon
Data Source
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AI summary
An object of the present invention is to provide a kit and a method capable of achieving high-precision measurement of a measurement target substance in a biological sample in a wide concentration range from a low concentration to a high concentration by sufficiently avoiding the influence of antibodies such as anti-serum albumin antibody present in blood. According to the present invention, there is provided a kit for measuring a measurement target substance in a biological sample, the kit including: a labeled particle having a first binding substance capable of binding to the measurement target substance and having a first blocking agent; and a substrate having a second binding substance capable of binding to any one of the measurement target substance or the first binding substance and having a second blocking agent, in which the labeled particle is a luminescent labeled particle containing at least one kind of compound represented by Formula (1) and a particle, and the first blocking agent and the second blocking agent are different from each other. Each symbol in Formula (1) has the meaning described in the present specification.