Homogeneous phase time discrimination fluorescence immunity analysis chelating agent and its preparing method

A time-resolved fluorescence and immunoassay technology, applied in chemical instruments and methods, luminescent materials, material testing products, etc., can solve the problems that the experimental application still has a certain distance, and the homogeneous immunoassay is limited to the laboratory stage.

Inactive Publication Date: 2008-07-16
CHANGCHUN INST OF APPLIED CHEMISTRY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The author believes that the research on homogeneous immunoassay is mostly limited to the laboratory stage, and there is still a certain distance from the experimental application.

Method used

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  • Homogeneous phase time discrimination fluorescence immunity analysis chelating agent and its preparing method
  • Homogeneous phase time discrimination fluorescence immunity analysis chelating agent and its preparing method
  • Homogeneous phase time discrimination fluorescence immunity analysis chelating agent and its preparing method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] (1) Synthesis of N-oxidized-2,6-dibromopyridine

[0043] Add 30.0g of 2,6-dibromopyridine, 180ml of trifluoroacetic acid, 40ml of hydrogen peroxide with a mass concentration of 30% to the reaction flask, stir and react at 35°C for 1h, cool to room temperature, pour into distilled water, and cool to -5 ℃, suction filtration, filtrate with anhydrous Na 2 CO 3 Neutralize to a pH value of 9, extract with chloroform, combine the organic phases, and evaporate the solvent to obtain a white solid product N-oxide-2,6-dibromopyridine.

[0044] (2) Synthesis of N-oxidation-2,6-dibromo-4-nitropyridine

[0045] Dissolve 19.8g of N-oxide-2,6-dibromopyridine in 67.8ml of concentrated H 2 SO 4 , add 67.8ml concentrated H under stirring 2 SO 4 And 34.2mL fuming nitric acid mixed acid, warming up to 80 ℃, stirring for 1h. After cooling, pour it into crushed ice, filter it with suction, and dry it in vacuo to obtain N-oxide-2,6-dibromo-4-nitropyridine as a light yellow solid produc...

Embodiment 2

[0063] (1) Synthesis of N-oxidation-2,6-dibromopyridine

[0064] Add 30.0g of 2,6-dibromopyridine, 190ml of trifluoroacetic acid, 40ml of hydrogen peroxide with a mass concentration of 30% to the reaction flask, stir and react at 38°C for 2h, cool to room temperature, pour into distilled water, and cool to -5°C , suction filtration, filtrate with anhydrous Na 2 CO 3 Neutralize to a pH value of 10, extract with chloroform, combine the organic phases, and evaporate the solvent to obtain a white solid product N-oxide-2,6-dibromopyridine.

[0065] (2) Synthesis of N-oxidation-2,6-dibromo-4-nitropyridine

[0066] Dissolve 19.8g of N-oxide-2,6-dibromopyridine in 67.8ml of concentrated H 2 SO 4 , add 67.8ml concentrated H under stirring 2 SO 4 And 34.2mL fuming nitric acid mixed acid, warming up to 90 ℃, stirring for 2h. After cooling, pour it into crushed ice, filter it with suction, and dry it in vacuo to obtain N-oxide-2,6-dibromo-4-nitropyridine as a light yellow solid pro...

Embodiment 3

[0084] (1) Synthesis of N-oxidized-2,6-dibromopyridine

[0085] Add 30.0g of 2,6-dibromopyridine, 200ml of trifluoroacetic acid, 40ml of hydrogen peroxide with a mass concentration of 30% to the reaction flask, stir and react at 42°C for 5h, cool to room temperature, pour into distilled water, and cool to -5 ℃, suction filtration, filtrate with anhydrous Na 2 CO 3 Neutralize to a pH value of 11, extract with chloroform, combine the organic phases, and evaporate the solvent to obtain a white solid product N-oxide-2,6-dibromopyridine.

[0086] (2) Synthesis of N-oxidation-2,6-dibromo-4-nitropyridine

[0087] Dissolve 19.8g of N-oxide-2,6-dibromopyridine in 67.8ml of concentrated H 2 SO 4 , add 67.8ml concentrated H under stirring 2 SO 4 And 34.2mL fuming nitric acid mixed acid, warming up to 100 ℃, stirring for 3h. After cooling, pour it into crushed ice, filter it with suction, and dry it in vacuo to obtain N-oxide-2,6-dibromo-4-nitropyridine as a light yellow solid prod...

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Abstract

The invention relates to a homogeneous time resolved fluorescence immunoassay chelating agent and the preparation method thereof. The chelating agent is N, N, N', N'-((2, 6-di(3'-aminomethyl)-1-pyrazole)-4-(4-isothiocyanate(phenylacetylene)-pyridine))-tetraacetic acid. A luminous group is 2, 6-di(pyrazole) pyridine; the invention has high triplet-state energy level, the excitation wavelength after the combination with rare earth ions is 320nm, the emission wavelength is 520 to 620nm, the luminous service life is 2.99ms, the quantum yield is 0.58, the invention is in line with the requirements of homogeneous TRFIA to be combined with phenylacetylene at the pyridine 4-position, and the acetylene bond can block protein macromolecules from consuming luminous energy. The phenyl 4-linking isothiocyanate at the pyridine 4-position can be coupled with protein without injury, so as to be conductive to the high-efficient measurement of the luminescence of the chelating agent. The polyammonia polycarboxyl structure of 11 coordination site can exclude the quenching function of water molecules on rare earth luminescence and improve the solubility of the chelating agent in water and the dynamic stability of the chelating agent in the water r solution.

Description

technical field [0001] The invention belongs to a homogeneous time-resolved fluorescent immunoassay chelating agent and a preparation method thereof. Background technique [0002] Homogeneous immunoassay (Homogeneous) is an analytical method that can be directly measured in liquid phase without repeated washing to separate bound and free markers. This method has long been one of the main goals pursued by immunoassay methodology research because of its simple operation, rapidity, and ease of automatic operation. [0003] Since Rubenstein reported the homogeneous enzyme labeling immunoassay method in 1972, more than 10,000 research reports on homogeneous and heterogeneous enzyme labeling analysis have proved that homogeneous analysis cannot exclude proteins, heme and various enzymes in plasma. The interference of it reduces its sensitivity, so it is mostly used for the analysis of certain drugs with higher concentrations in body fluids. (Reference Ngo.TTEnzyme-Mediated Immun...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/52G01N33/58C09K11/06
Inventor 潘利华马世盐谢文兵钮倩石竹李珍刘云霞
Owner CHANGCHUN INST OF APPLIED CHEMISTRY - CHINESE ACAD OF SCI
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