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A Method for Judging Hook Effect in Homogeneous Time-Resolved Fluorescence Immunoassay

A technology of time-resolved fluorescence and hook effect, applied in the direction of material analysis, analysis of materials, fluorescence/phosphorescence, etc. by optical means, it can solve the problems of affecting detection throughput, increase cost, consumption, etc., and achieve high detection throughput. Effect

Active Publication Date: 2021-11-02
SUZHOU INST OF BIOMEDICAL ENG & TECH CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Disadvantages: Adding target substances brings additional reagent consumption and increases costs; it can only be judged at the end of the reaction
At present, only Thermo Fisher's Kryptor series products are used clinically abroad, and its patent for judging the hook effect has not yet been retrieved. According to public information, it uses the reaction kinetic curve within 60 seconds in the initial stage of the immune reaction to judge the sample. If the concentration is too high and needs to be diluted, it needs to be read at 9 time points, which will cause the detection system to be occupied for a long time and affect the detection throughput

Method used

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  • A Method for Judging Hook Effect in Homogeneous Time-Resolved Fluorescence Immunoassay
  • A Method for Judging Hook Effect in Homogeneous Time-Resolved Fluorescence Immunoassay
  • A Method for Judging Hook Effect in Homogeneous Time-Resolved Fluorescence Immunoassay

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1: Hook effect assay for AFP assay

[0043] 2.3 Generate Calibration Curve

[0044] Detect the signal absolute value of a series of known concentration samples, and use the relationship between the measured signal absolute value and rate value and concentration to establish two calibration curves, within the range of positive correlation between concentration and signal (see figure 2 ), using four parameters for fitting to generate a calibration curve, the fitting curve of the relationship between the absolute value of the signal and the concentration is shown in image 3 , the fitting curve between the rate value and the concentration is shown in Figure 4 .

[0045] 2.4 Calculate the concentration of unknown samples using the calibration curve

[0046] Using the fitted four-parameter curve equation, the concentration value of the unknown sample can be calculated, and using two curves, two concentration values ​​can be calculated.

[0047] Concentr...

Embodiment 2

[0053] Example 2: Hook effect detection for AFP assays across different systems

[0054] 2.3 Generate Calibration Curve

[0055] The signal value and rate value measured between different systems (instruments) are different, Figure 5 Shows the systematic deviation of signal values ​​between the two instruments. Compared with Example 1, Example 2 introduces the 665 / 620nm measurement ratio of the blank sample as the background value, and the measurement system deviation of the calibration signal between different systems (instruments), the calculation method is as follows: divide the signal absolute value and the rate value With the background value, the relative signal absolute value (RS, relative signal) and the relative rate value (RRR, relative reaction rate) were respectively obtained. Under the same incubation conditions, the RS and RRR curves are only related to the degree of immune response, and have nothing to do with the deviation of the measurement system. Using th...

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Abstract

The invention relates to a method for judging the hook effect of homogeneous time-resolved fluorescence immunoassay. The fluorescence intensity ratio of two wavelengths is used as the signal value; (2) by measuring the signal values ​​at least two moments in the initial stage of the reaction, by linear fitting and four-parameter fitting, the absolute value of the signal intensity and the rate value of the signal change are respectively established. and the standard curve of the sample concentration; (3) detect the signal value of the unknown sample, and obtain the concentration of the unknown sample according to the fluorescence intensity of the unknown sample through the standard curve, thereby avoiding the hook effect. Using the ratio of the absolute value of the signal intensity and the value of the rate of change of the signal to the background signal, the relative signal intensity and the relative value of the rate of change of the signal are calculated to achieve consistent results on different instruments. The concentration interpretation of more samples can be completed within a given time, so that the system has higher detection throughput.

Description

technical field [0001] The invention relates to a method for judging the hook effect of homogeneous time-resolved fluorescence immunoassay, belonging to the technical field of fluorescence immunoassay. Background technique [0002] The homogeneous time-resolved fluorescence immunoassay technique combines two techniques of fluorescence resonance energy transfer (FRET, Fluorescence Resonance Energy Transfer) and time-resolved fluorescence (TRF, Time Resolved Fluorescence). This technology utilizes chelate markers of rare earth elements with a crypt structure as fluorescent donors, and short-lived fluorescent molecules with good spectral overlap with the fluorescent donors as fluorescent acceptors. Fluorescence resonance energy transfer (FRET) occurs with the acceptor (second fluorescent label). In fluorescence resonance energy transfer, the lifetime of the acceptor to emit fluorescence is equal to the lifetime of the donor to emit fluorescence. Because the donor fluorescence...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/64G01N33/53G01N1/38
CPCG01N21/6408G01N21/6402G01N33/53G01N1/38
Inventor 刘涛
Owner SUZHOU INST OF BIOMEDICAL ENG & TECH CHINESE ACADEMY OF SCI
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