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Application of indole derivative as chemiluminescence enhancer and kit containing enhancer

A chemiluminescence reagent and chemiluminescence technology, applied in biological testing, scientific instruments, instruments, etc., can solve the problems of large increase in brightness and poor stability

Pending Publication Date: 2022-07-15
大连博格林生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, the existing phenothiazine enhancers and the second enhancers that have been reported have a large increase in brightness, but poor stability

Method used

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  • Application of indole derivative as chemiluminescence enhancer and kit containing enhancer
  • Application of indole derivative as chemiluminescence enhancer and kit containing enhancer
  • Application of indole derivative as chemiluminescence enhancer and kit containing enhancer

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0090] In this example, formula 1 is used for antigen-antibody detection.

[0091] This kit can be used for the detection of various types of antibodies and antigens in vitro. Antigens and antibodies are coated in white or black opaque polystyrene microplates, and then the content of antigens or antibodies is detected by the conjugated HRP marker. Take alpha-fetoprotein (AFP) for example.

[0092] The reference substance is a commercially available alpha-fetoprotein quantitative detection kit (chemiluminescence immunoassay (Transview Life), using the reagents used in the calibrator for detection and establishment of a standard curve, coated in a white polystyrene microplate, The concentration range is 0-500ng / ml. And use the corresponding antibody conjugated to HRP to incubate, use the BioTek Synergy H1 full-function microplate detector to detect its luminescence intensity, and select the microwell with a concentration of 250ng / ml to detect the luminescence once every 5min va...

Embodiment 2

[0098] In this implementation, formula 2 is used to compare the thermal stability of chemiluminescence reagents.

[0099] The reagents were prepared according to formula 2, and the obtained reagents were placed at 37 degrees Celsius, 4 days, 8 days, and 12 days, respectively, and compared with the preparation of the luminescent reagent on the same day. The comparison experiment directly used horseradish peroxidase dissolved in physiological saline, Dilute root peroxidase to a concentration of 1X10 -15 mol / L for later use, take 100ul of each of the luminescent solutions A and B that have been stored for different days, add them to a white polystyrene microplate after mixing, and then add the diluted horseradish peroxidase to each well. The luminescence value was detected using a BioTek Synergy H1 full-function microplate reader. The results are shown in Table 2.

[0100] Table 2 Thermal stability test of chemiluminescent reagents

[0101]

[0102] According to the above r...

Embodiment 3

[0104] Example using formula three, applied to western blot detection detection effect

[0105] The specific operation process is as follows:

[0106] Select mouse liver tissue, use RIPA lysis solution to extract protein, and then detect the protein concentration. According to the measured protein concentration, calculate the mass of protein added to each well of protein electrophoresis to be 100ug, 50ug, 25ug, 12.5ug, 6.25ug and 3.125ug respectively. . Protein electrophoresis and membrane transfer were performed according to routine operations. The membrane used was NC membrane. The NC membrane was blocked with nonfat milk powder, incubated with Histone H3 primary antibody overnight, and then incubated with HRP-labeled secondary antibody, and then detected by luminescence reagent. The selected control product is an imported best-selling product: SuperSignal West Dura chemiluminescent substrate. In order to ensure consistent results, the same NC membrane was used in the dete...

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Abstract

The invention belongs to the field of chemiluminescence reagents, and discloses application of an indole derivative as a chemiluminescence enhancer and a kit containing the enhancer. In the application, the indole derivative is used as a second reinforcing agent and is combined with a phenothiazine compound as a first reinforcing agent; furthermore, the phenothiazine compound is selected from one of 3-(10-phenothiazinyl) propane-1-sodium sulfonate, 10-(3-dimethyl aminopropyl) phenothiazine and derivatives of the 10-(3-dimethyl aminopropyl) phenothiazine; furthermore, the indole derivative is selected from one or more of tryptophan, 5-hydroxytryptamine, auxin or indole-3-acetic acid. The phenothiazine compound and the indole derivative are combined for use to play a synergistic role, so that the brightness of the luminescent reagent is improved, and meanwhile, the luminescence time is prolonged due to mutual transmission of electrons.

Description

technical field [0001] The invention belongs to the field of chemiluminescence reagents, and discloses the use of indole derivatives as chemiluminescence enhancers and a kit containing the enhancers. Background technique [0002] Horseradish peroxidase (HRP)-catalyzed chemiluminescence reaction of luminol and peroxygen compounds is widely used in the detection and analysis of antigens and antibodies, especially in enzyme-linked immunosorbent assay (ELISA) and western blotting experiments. Horseradish peroxidase is coupled to the antigen or antibody, and the amount of antigen or antibody is indirectly analyzed by measuring the amount of HRP. [0003] As a classic chemiluminescence reaction, luminol has been successfully used to enhance HRP-induced chemiluminescence, including: p-iodophenol, p-coumarin and other phenolic enhancers, in addition to phenolic enhancers, there are acetyl Anilines, phenothiazines, of which phenothiazines belong to the newly discovered enhancers. P...

Claims

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Application Information

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IPC IPC(8): G01N33/53G01N33/532
CPCG01N33/5306G01N33/532
Inventor 李广亮李民强
Owner 大连博格林生物科技有限公司
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