Carcino-embryonic antigen immunization analysis method based on singlet oxygen channel light-emitting quantum dot sensor

A carcinoembryonic antigen and immunoassay technology, applied in the field of immunodetection, can solve the problems of near-infrared sensitive detector requirements still existing, no more than 10nm, spectral width, etc.

Inactive Publication Date: 2016-11-09
SOUTHERN MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Moreover, the FRET transmission distance is limited, and generally cannot exceed the spatial distance of 10nm
On the other hand, due to the broad fluorescence emission spectrum of most organic dyes and obvious tailing, the existing HTRFA technology is only suitable for analysis in the long-wavelength or near-infrared band, and the requirement for expensive near-infrared sensitive detectors still exists

Method used

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  • Carcino-embryonic antigen immunization analysis method based on singlet oxygen channel light-emitting quantum dot sensor
  • Carcino-embryonic antigen immunization analysis method based on singlet oxygen channel light-emitting quantum dot sensor
  • Carcino-embryonic antigen immunization analysis method based on singlet oxygen channel light-emitting quantum dot sensor

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Embodiment 1

[0051] The core technology of this step is to obtain symmetrical water-soluble quantum dots with high quantum yield and narrow maximum half-maximum width of the fluorescence emission peak. The concrete implementation of the present invention used is the core / shell type cadmium selenide / zinc sulfide quantum dot purchased from U.S. Invitrogen Company, and its solvent is decane, and its spectral Stock shift length is from 592nm to the wider range between short wavelength Therefore, the quantum dot can be excited with a wavelength shorter than 592nm; the fluorescence emission peak is 605±0.5nm, the maximum half-peak width is 27±1.5nm, and the quantum yield is 76%; Oxygen-receiving "antenna" thiazine compound has a matching fluorescence emission spectrum, overcomes the shortcomings of organic dyes in all aspects, and is suitable for replacement to reduce measurement errors, improve fluorescence resonance energy transfer (FRET) efficiency, and improve analytical sensitivity.

[0052...

Embodiment 2

[0062] A carcinoembryonic antigen immunoassay method based on singlet oxygen channel luminescence quantum dot sensor, comprising quantum dot nano hydrosol and functional photosensitive microsphere; the preparation method of quantum dot nano hydrosol and functional photosensitive microsphere is as follows:

[0063] 1. Preparation of high-performance water-soluble quantum dots: put 50 μL of fat-soluble quantum dots in a centrifuge tube, add 200 μL of anhydrous methanol; mix well, seal the centrifuge tube, centrifuge at 3000 rpm, 4 ° C for 5 min; discard the supernatant Add 40 μL of chloroform to fully dissolve; add 21 mg of glutathione stabilizer and 10 μL of 0.1M sodium hydroxide aqueous solution, mix well, shake and incubate on a shaker, and store at room temperature for 2 hours; add 100 μL of deionized water and 1.8 mL of acetone, precipitated, centrifuged at 6000 rpm for 5 min; dissolved in 0.5 mL of deionized water to obtain water-soluble quantum dots. The emission spectrum...

Embodiment 3

[0068] Based on the application of singlet oxygen channel luminescent quantum dot sensor in the analysis of homogeneous oxygen transfer fluorescence resonance energy transfer, the principle of homogeneous analysis of singlet oxygen channel luminescent quantum dot sensor is as follows image 3 As shown, the steps to apply are as follows:

[0069] 1. Conjugate TQPS with anti-carcinoembryonic antigen monoclonal antibody, the steps are as follows: prepare 0.5M MES buffer, adjust pH to 6.1, take 100 μL for use; prepare 50 mg / mL Sulfo-NHS solution with deionized water, take 230 μL Stand-by; prepare 50 mg / mL EDAC solution with deionized water, take 240 μL for use; mix the above solution for use, dilute to 1 mL with deionized water, add to the TQPS prepared in the above step 3, and resuspend; the mixture is at room temperature Shake the reaction for 30 minutes; centrifuge at 16000rmp to remove unreacted Sulfo-NHS and EDAC, wash with deionized water, resuspend, and centrifuge three tim...

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Abstract

The invention discloses a carcino-embryonic antigen immunization analysis method based on a singlet oxygen channel light-emitting quantum dot sensor. The method is characterized in that the quantum dot nano hydrosol and a monoclonal antibody 1 of to-be-measured carcino-embryonic antigen are coupled, the functional photosensitive microspheres and streptavidin are coupled, a monoclonal antibody 2 of to-be-measured carcino-embryonic antigen is subjected to biotin labeling, a double-antibody sandwich method is used, through irradiation of the near infrared laser with 680 nm, a fluorescence signal at 605 nm is collected, the intensity of the fluorescence signal is proportional to the content of carcino-embryonic antigen in a system, so that the carcino-embryonic antigen homogeneous fluoroimmunoassay is realized. The carcino-embryonic antigen immunization analysis method is established based on the singlet oxygen channel light-emitting quantum dot sensor, and the thereby usage of an organic dye as a fluorescent receptor is avoided. The detection method has the advantages of high sensitivity, good stability, and simple and convenient usage.

Description

technical field [0001] The invention relates to the technical field of immune detection, in particular to an immunoassay method for carcinoembryonic antigen based on a singlet oxygen channel luminescent quantum dot sensor. Background technique [0002] Carcinoembryonic antigen (CEA) is a glycoprotein complex with a molecular weight of about 220KDa. It was first discovered in colon cancer tissue extracts by Canadian scientists Samuel O. Freedman and Phil Gold in 1965. Generally speaking, carcinoembryonic antigen is synthesized by gastrointestinal tract tissue, pancreas and liver cells during fetal period, and basically ceases to be synthesized after birth. Therefore, the content of carcinoembryonic antigen in the serum of normal healthy individuals is maintained at a very low level, about 2.5 μg / mL. It is generally believed that elevated levels of CEA in serum are more common in patients with gastric cancer, pancreatic cancer, lung cancer, breast cancer, and medullary thyroi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577G01N21/64
CPCG01N33/577G01N21/6428
Inventor 刘天才陈振华
Owner SOUTHERN MEDICAL UNIVERSITY
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