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Quantum dot fluorescent probe and application thereof
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A technology of fluorescent probes and quantum dots, which is applied in the field of quantum dot fluorescent probes to achieve the effects of high yield, strong fluorescent signal and highly sensitive detection
Inactive Publication Date: 2015-12-16
SHANGHAI INST OF MICROSYSTEM & INFORMATION TECH CHINESE ACAD OF SCI +1
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Embodiment 1
[0023] Example 1: Fluorescent quantum dot probe capable of detecting a protein carcinoembryonicantigenCEA
[0024] (1) Preparation of quantum dot fluorescent probes
[0025] First, take 10-50μL of 5-50mM SMCC to activate 50-500μL of amino quantum dots 625 (Qdot625), 25°C, 30-60min; then take the method of desalting column for purification; use 10-50mM DTT to 100-500μL CEA detection antibody Reduction, 25°C, 30-60min; after the desalting column removes unreacted antibodies and other impurities, incubate it with the purified activated quantum dots in a hybridizer, 25°C, 30-60min; add 5-50μL β-thiol The reaction was terminated with ethanol, and after separation and purification, it was stored in 10 mM PBS (PH7.2) at 4°C in the dark for use.
[0026] (2) Detection reaction
[0027]Add 5000-10000 in vitro CEA capture antibody-labeled magnetic bead capture probes and 5-50nMQdot625 detection probes to each reaction, and make up the volume to 10-50ul / reaction, then add 10-50ul of d...
Embodiment 2
[0030] Example 2: Fluorescent quantum dot probe capable of detecting various proteins CEA, neuron-specific enolase (NSE), and cytokeratin fragment 19 (CYFRA21-1)
[0031] (1) Preparation of quantum dot fluorescent probes
[0032] First, take 10-50μL of 5-50mM SMCC to activate 50-500μL of amino quantum dots Qdot525, Qdot585, Qdot625, 25℃, 30-60min; then use desalting column method to purify; use 10-50mM DTT to 100-500μL NSE, CEA , CYFRA21-1 detection antibody for reduction, 25°C, 30-60min; after desalting column to remove unreacted antibody and other impurities, incubate it with purified activated quantum dots in a hybridizer, 25°C, 30-60min ; Add 5-50 μL of β-mercaptoethanol to terminate the reaction, and store in 10 mM PBS (PH7.2) after separation and purification, at 4°C in the dark for later use.
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Abstract
The invention relates to a quantum dot fluorescent probe and application thereof. The invention is characterized in that the quantum dot fluorescent probe utilizes a difunctional cross-linking agent long-chain succinimidyl-4-[N-maieimidomethyl]cyclohexane-1-carboxy-[6-amidocaproate] (SMCC) to undergo a cross-linking reaction with an amino group on the surface of an amino quantum dot; a maleimide group is produced on the surface of the quantum dot; a to-be-labeled antibody is reduced by a reducing agentdithiothreitol (DTT) so as to reduce a disulfide bond into a mercapto group; and the mercapto group and the maleimide group form a covalent bond so as to realize labeling of the quantum dot with the antibody. The quantum dot fluorescent probe is applied to (1) detection of one protein oncofetal antigen or (2) detection of a plurality of protein CEA, euron-specific enolase (NSA) and a cytokerain fragment 19CYFRA21-1, wherein the detection limit of detection (1) is 38 pg / ml, and the detection limit of detection (2) is 0.9 ng / ml.
Description
technical field [0001] The invention relates to the technical field of nano-biological detection, in particular to a quantum dot fluorescent probe, which can simultaneously detect multiple biological molecules with high sensitivity. Background technique [0002] Being able to detect biomolecules with high sensitivity and high specificity has always been the direction of people's efforts, and it is of great significance in the fields of early detection of diseases, rapid diagnosis, health detection, and environmental monitoring. [0003] Quantum dots, also known as semiconductor nanocrystals or semiconductor nanoparticles, are nanoparticles composed of II-VI group (such as cesium cadmium CdSe, cadmiumtelluride CdTe) or III-V group (such as InP, InAs) elements. The particle size is generally 1-10 nm. Since electrons and holes are quantum confined, the continuous energy band structure becomes a discrete energy level structure with molecular characteristics, which can emit fluo...
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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68G01N33/533
CPCG01N33/533G01N33/54393G01N33/68
Inventor 贾春平吴思敏李宫刘莉芬郜晚蕾景奉香金庆辉赵建龙
Owner SHANGHAI INST OF MICROSYSTEM & INFORMATION TECH CHINESE ACAD OF SCI
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