Enzymatic Chemiluminescent Substrates

An enzymatic chemiluminescence, substrate technology, applied in the field of immunodetection, can solve the problems of long-term stable preservation, limited application by storage conditions and use conditions, poor stability, etc., to improve thermal stability, real-time stability, duration The effect of long and fast entry into the plateau

Active Publication Date: 2021-01-01
普十生物科技(北京)有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In related technologies, APS-5 enzymatic chemiluminescent substrates have poor stability and cannot be stored stably for a long time. The difficulty of their storage and use conditions limits their application to a certain extent.

Method used

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  • Enzymatic Chemiluminescent Substrates
  • Enzymatic Chemiluminescent Substrates
  • Enzymatic Chemiluminescent Substrates

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Preparation of Enzymatic Luminescent Substrates

[0031] (1) Material: 5-(tetradecamido)fluorescein, 9-(4-chlorophenylthiobenzoyloxymethylene)-10-methyl-9,10-dihydroacridine-disodium Salt, MgCl 2 , ZnCl 2 , aryl acylhydrazone-9,10 acridine derivatives, cetyl trimethyl ammonium chloride are commercially available, chemically pure; Proclin-300, Tween20, Tris are products of sigma-aldrich in the United States. Among them, aryl acylhydrazone-9,10 acridine derivatives are synthesized from aryl acyl hydrazones and 9,10 acridine derivatives.

[0032] (2) Using Tris-Tween20 buffer as a solvent, different concentrations of enzymatic chemiluminescence substrates were prepared, as shown in Table 1.1-Table 1.4.

[0033] Table 1.1: Recipe one

[0034]

[0035] Table 1.2: Recipe II

[0036]

[0037]

[0038] Table 1.3: Recipe three

[0039]

[0040] Table 1.4: Recipe Four

[0041]

[0042]

[0043] In the enzymatic chemiluminescence substrate provided by the ...

Embodiment 2

[0045] Comparing the enzymatic chemiluminescence substrates of formula 1, formula 2, formula 3 and formula 4 with the commercially available Lumigen luminescent substrate, the AP signal intensity and linear relationship of each gradient concentration were tested. The details are shown in Table 2.1-Table 2.5:

[0046] Table 2.1

[0047]

[0048] Table 2.2

[0049]

[0050] Table 2.3

[0051]

[0052] Table 2.4

[0053]

[0054] Table 2.5

[0055]

[0056] From the detection data results in Table 2.1-Table 2.5, it can be seen that the blank luminescence intensity of the enzymatic chemiluminescence substrate provided by the present invention is not higher than 300; under the condition of the same concentration of AP, the enzymatic chemiluminescence substrate provided by the present invention The signal intensity is higher than that of the commercially available Lumigen luminescent substrate.

Embodiment 3

[0058] The linearity of the enzymatic chemiluminescence substrates of formula 1, formula 2, formula 3 and formula 4 was compared with that of the commercially available Lumigen luminescent substrates for the determination of carcinoembryonic antigen (CEA) quantitative detection kits.

[0059] The double antibody sandwich method was used to detect the content of CEA in human serum. The experimental steps were as follows:

[0060] Step S1: Add 30ul of human serum sample or CEA calibrator, 100ul of diluted enzyme-labeled antibody, and 30ul of antibody-conjugated magnetic beads into the chemiluminescence tube, respectively, and react at 37°C for 15min after mixing;

[0061] Step S2: add 300ul of cleaning solution to clean 3 times;

[0062]Step S3: adding 100ul of the enzymatic chemiluminescence substrates of formula 1, formula 2, formula 3, formula 4 and commercially available chemiluminescence substrates of Lumigen into different chemiluminescence tubes respectively;

[0063] St...

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Abstract

The invention discloses an enzymatic chemiluminescent substrate. The enzymatic chemiluminescent substrate comprises the following components: 9-(4-chlorphenyl thio-benxoyl oxymethylene)-10-methyl-9-10-acridanyl-disodium salt, 5-(tetradecanamide) fluorescein and an aryl acylhydrazone-9, 10 acridine derivate. According to the enzymatic chemiluminescent substrate, the aryl acylhydrazone-9, 10 acridine derivate is added as a substrate enhancer which is dissolved in a substrate solution and can act with alkaline phosphatase to light, so that the enzymatic chemiluminescent substrate has the advantages of being high in lighting signal, fast to enter plateau, long in lasting time, and long in normal-temperature storage term.

Description

【Technical field】 [0001] The invention relates to the technical field of immunodetection, in particular to an enzymatic chemiluminescence substrate. 【Background technique】 [0002] Chemiluminescence is a phenomenon of light radiation accompanying substances in the process of chemical reaction, which can be divided into direct luminescence and indirect luminescence. Direct luminescence is the simplest chemiluminescence reaction and consists of two key steps: excitation and radiation. For example, two substances A and B react chemically to form C substance. The energy released by the reaction is absorbed by the molecules of C substance and transitions to the excited state C*, and the excited C* generates light radiation in the process of returning to the ground state. Here C* is a luminophore. In this process, C is directly involved in the reaction, so it is called direct chemiluminescence. Indirect luminescence, also known as energy transfer chemiluminescence, is mainly com...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/76G01N33/532
CPCG01N21/76G01N33/532
Inventor 王琳康丽波
Owner 普十生物科技(北京)有限公司
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