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Competitive homogeneous chemiluminescence detection method and application thereof

A homogeneous chemiluminescence and detection method technology, which is applied in the direction of chemiluminescence/bioluminescence, analysis by making materials undergo chemical reactions, and measurement devices, can solve the problems of not being able to effectively balance functional sensitivity and analysis range, and achieve broadened detection Effects of range, prevention of hook effects, high precision and accuracy

Pending Publication Date: 2021-07-16
CHEMCLIN DIAGNOSTICS CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, for some special indicators such as steroid hormones, there are high requirements for functional sensitivity and detection range, and existing chemiluminescence immunoassays, electrochemiluminescence immunoassays, photochemiluminescence immunoassays, etc. still have defects , cannot effectively take into account the special requirements of functional sensitivity and analytical range

Method used

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  • Competitive homogeneous chemiluminescence detection method and application thereof
  • Competitive homogeneous chemiluminescence detection method and application thereof
  • Competitive homogeneous chemiluminescence detection method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0088] Embodiment 1: the preparation of the reagent used for the method of the present invention that the analyte is testosterone Reagent and instrument:

[0089] Testosterone Antibody Conjugated to Biotin, Receptor Microspheres, BSAylated Dihydrotestosterone (BSA-DHT), BSAylated Testosterone (BSA-T), Phosphate Buffer (0.02M PBS, pH 7.2), BSA, Tween-20, LiCA 500 (Beijing Kemei Biotechnology Co., Ltd.), Hitachi high-speed refrigerated centrifuge.

[0090] making process:

[0091] (1) Preparation of competitive antigen-coupled receptor microsphere solution (reagent 1)

[0092] 1) Take 2 mg of receptor microspheres from two 2 mL centrifuge tubes, centrifuge at 10,000 rpm at 4°C for 15 min, and wash once.

[0093] 2) Ultrasonic dispersion is uniform, add 0.2mg BSA-T to one of the centrifuge tubes, add 0.02mg BSA-DHT to the other, mix well and coat overnight at 4°C;

[0094] 3) Add 20uL 10mg / mL BSA-blocked receptor microspheres to two centrifuge tubes, and rotate at room tempera...

Embodiment 2

[0104] Utilize R1-1, R1-2, R1-3 prepared in Example 1 respectively as the method of reagent 1 to detect the sample containing testosterone, and the detection result is compared with the Beckman measured value, and the results are respectively as follows Figure 2-4 shown.

[0105] The detection process is automatically completed by the LiCA500 automatic light-induced chemiluminescence analysis system and the detection results are output. The specific steps are:

[0106] a. Add 20 μl of sample, calibrator or quality control to the reaction well;

[0107] b. Add 20 μl release agent, 25 μl reagent 1 and 25 μl reagent 2 to the reaction well in sequence;

[0108] c. Incubate at 37°C for 15 minutes;

[0109] d. Add 175 μl of LiCA universal solution (donor microsphere solution combined with avidin);

[0110] e. Incubate at 37°C for 15 minutes;

[0111] e. Laser irradiates microholes and calculates the amount of photons emitted per hole;

[0112] f. According to the calibration c...

Embodiment 3

[0114] Embodiment 3: precision detection

[0115] Intra-batch precision detection: use the reagents, high, middle and low samples adopted by the method of the present invention to carry out precision detection: every batch of reagents is measured 10 times, and the mean value X and standard deviation SD of 10 measurement results are calculated, according to the formula CV=SD / X×100% to get the coefficient of variation CV. The results are shown in Table 1 and Table 3.

[0116] Inter-batch precision detection: use three batches of reagents used in the method of the present invention to detect high, middle and low samples respectively, repeat 10 times, calculate the mean value X and standard deviation SD of 30 measurement results, according to the formula CV=SD / X× 100% yields a coefficient of variation, CV. The results are shown in Table 2 and Table 3.

[0117] Table 1: Raw data of intra-assay precision of LiCA T reagent (photo-induced chemiluminescence method)

[0118]

[...

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Abstract

The invention relates to a competitive homogeneous chemiluminescence detection method. The competitive homogeneous chemiluminescence detection method comprises the following steps: contacting an analyte with a first composition, a reagent containing a second composition and a third composition and a fourth composition to form a mixture to be detected; providing exciting light to irradiate the mixture to be detected at least once; and then detecting the intensity of the generated chemiluminescence signal so as to judge whether the analyte exists or not and / or the concentration of the analyte. According to the method, ''differential receptor microspheres'' are utilized so that the method has excellent functional sensitivity and a detection range at the same time.

Description

technical field [0001] The invention belongs to the technical field of homogeneous chemiluminescence, and in particular relates to a competitive homogeneous chemiluminescence detection method and its application. Background technique [0002] The competitive immunoassay is an assay for the quantitative analysis of small molecule haptens. Radioimmunoassay (RIA) is the earliest established competitive immunoassay method and won the 1974 Nobel Prize in Physiology and Medicine. In the radioimmunoassay, the competing antigen (labeled antigen) containing radionuclide labeling and a limited amount of specific antibody, the antigen to be tested in the specimen and the labeled antigen as a reagent compete with the specific antibody respectively. Separate the bound marker (B) and the free marker (F), and measure the radioactivity (or intensity, in counts per minute, CPM) of the bound marker. The radioactivity is inversely proportional to the antigen to be tested. A series of calibra...

Claims

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Application Information

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IPC IPC(8): G01N21/76G01N33/543G01N33/577
CPCG01N21/76G01N33/54313G01N33/577
Inventor 范树芹强中华徐静心李临
Owner CHEMCLIN DIAGNOSTICS CO LTD
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