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Synthesis and application of fluorescent probe with specific recognition on GC33-3-1 antibody

A fluorescent probe and specific technology, applied in fluorescence/phosphorescence, luminescent materials, organic chemistry, etc., can solve the problems of affecting drug clearance rate, affecting the apparent distribution volume of drugs, etc., achieving simple synthesis method, simple post-processing, The effect of easy availability of raw materials

Inactive Publication Date: 2018-11-23
FUJIAN MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In most cases, the interaction between protein and drug will significantly affect the apparent volume of distribution of the drug and also affect the clearance rate of the drug

Method used

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  • Synthesis and application of fluorescent probe with specific recognition on GC33-3-1 antibody
  • Synthesis and application of fluorescent probe with specific recognition on GC33-3-1 antibody
  • Synthesis and application of fluorescent probe with specific recognition on GC33-3-1 antibody

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Example 1: Synthesis of small molecule fluorescent probes for antibody labeling

[0038] (1) Synthesis of intermediate compound F1:

[0039]

[0040] Weigh 15.7 mmol of 4,6-dichloropyrimidine and o-phenylenediamine, and 17.5 mmol of N,N-diisopropylethylamine in a microwave reaction tube, add 10 ml of isopropanol to dissolve, and react in the microwave at 150°C for 10 min. Post-processing: TLC was used to monitor the reaction results, and the reaction solution was evaporated under reduced pressure to remove the solvent to obtain yellow oil F1 with a yield of 100%, MS (m / z): 220.5.

[0041] (2) Synthesis of intermediate compound F2:

[0042]

[0043] Weigh 1.6 mmol of F1, 4.4 mmol of carbonyl-2--imidazole, and 4.4 mmol of pyridine into a round bottom flask, add tetrahydrofuran as a solvent, protect with nitrogen, and reflux in an oil bath at 65°C overnight. Post-processing: TLC was used to monitor the reaction results, the reaction solution was evaporated under r...

Embodiment 2

[0047] Example 2: Fluorescence changes after fluorescent probe-labeled human serum albumin (HSA) reaction

[0048] Sample solution: a certain amount of fluorescent probe prepared in Example 1 was dissolved in an appropriate amount of methanol solution and diluted with ultrapure water to a concentration of 1.5×10 -5 mol / L; Dilute human serum albumin (HSA) with ultrapure water to a concentration of 1X10 -5 mol / L; Tris-HCl solution is prepared to be 0.05mol / LPH=7.4 (contains 0.1mol / L NaCl); add 1ml Tris-HCl solution, 1ml fluorescent compound and 1 ml HSA solution, mix well. Control solution: 1 ml fluorescent probe solution, 1 ml Tris-HCl solution, and 1 ml ultrapure water were mixed to prepare a control solution. Determination of fluorescence spectrum image 3 .

Embodiment 3

[0049]Example 3: Fluorescent probes directly recognize antibodies

[0050] (1) Coating: Dilute the antibody to 1 µg / ml, mix it with the coating solution (carbonate buffer) and add it to the enzyme-labeled plate, incubate at 37°C for 2 hours, and overnight at 4°C;

[0051] (2) Washing: Take 40 ml PBS and add 20 µl Tween 20 to make PBST solution, prepare PBST solution, wash the enzyme-labeled plate with PBST solution, 300 µl per well, 3 min each time, wash 3 times;

[0052] (3) Blocking: Bovine serum albumin was prepared to 0.02 mg / ml, added to the microtiter plate, and incubated at 37°C for 2 h;

[0053] (4) Washing: wash the enzyme-labeled plate with PBST solution, 300 µl per well, 3 min each time, and wash 3 times;

[0054] (5) Binding: Dissolve a certain amount of the fluorescent probe compound prepared in Example 1 in an appropriate amount of methanol solution and dilute it with ultrapure water to a concentration of 1.5X10 -5 mol / L; Tris-HCl solution was prepared to be 0...

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Abstract

The invention discloses synthesis and application of a fluorescent probe with specific recognition on a GC33-3-1 antibody. A synthesis route of the probe is simple, reaction conditions are moderate and post-treatment is simple and convenient. After the fluorescent probe is labeled with human serum albumin (HSA), the fluorescence intensity is greatly enhanced; after the fluorescent probe is labeledwith the HSA, the fluorescent probe is used for recognizing the GC33-3-1 antibody. Compared with other methods for detecting the antibody, the method has the advantages of simplicity in operation, sensitive response, selectivity and the like, and has a relatively good application prospect in the field of biology; a formula is shown in the description.

Description

technical field [0001] The invention belongs to the field of biological analysis and detection, and in particular relates to the synthesis and application of a fluorescent probe for specifically recognizing GC33-3-1 antibodies. Background technique [0002] Human serum albumin (HSA) is the most abundant protein in serum and the most important drug carrier protein. It is involved in the transport, distribution and metabolism of many endogenous and exogenous lipids, including fatty acids, amino acids, metal ions and many drugs, and plays an important role in the mixing osmotic pressure of blood. Human serum albumin (HSA) is a high molecular weight plasma protein. It is a single chain of 585 amino acids. Like other mammalian albumins, human albumin contains 17 disulfide bridges and one free thiol Cys34. In most cases, the interaction between protein and drug will significantly affect the apparent volume of distribution of the drug and also affect the clearance rate of the dr...

Claims

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Application Information

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IPC IPC(8): C07D401/14C09K11/06G01N21/64
CPCC07D401/14C09K11/06C09K2211/1029C09K2211/1044G01N21/6428
Inventor 柯方巫小文许建华林巧发涂书清陈艳王津
Owner FUJIAN MEDICAL UNIV
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