Immunofluorescent double staining kit for cervical cancer auxiliary diagnosis

An auxiliary diagnosis and immunofluorescence technology, applied in the field of liquid-based cytology immunofluorescence staining, can solve the problems of low sensitivity and complexity, and achieve the effect of efficient excitation and low background.

Active Publication Date: 2020-05-01
江苏美克医学技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This technology has the characteristics of simplicity, high specificity, low sensitivity, and complicated detection of multiple antigens at the same time.
[0006] However, ordinary fluorescence double-staining kits need to excite different colors of fluorescence under different excitation lights, and then form double-stained images through image processing, which requires different fluorescence excitation modules and image processing systems.

Method used

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  • Immunofluorescent double staining kit for cervical cancer auxiliary diagnosis
  • Immunofluorescent double staining kit for cervical cancer auxiliary diagnosis
  • Immunofluorescent double staining kit for cervical cancer auxiliary diagnosis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] 1. Monoclonal antibody-labeled dextran amplification process:

[0038] (1) Dissolve dextran (DeX) in 20mM phosphate buffered saline (PBS) with a pH value of 7.0, add sodium periodate (NaIO 4 ) was oxidized, and the reaction was stirred at 30°C to 37°C in the dark for 2-3 hours and then terminated by adding excess ethylene glycol; the same PBS as the DeX matrix was used for dialysis overnight.

[0039] (2) Dialyze the required antibody against PBS overnight.

[0040] (3) After the oxidized DeX and the antibody were mixed and reacted, an appropriate amount of sodium cyanoborohydride (NaCNBH 3 ) The reaction time in the dark is 1.5-3 hours, and the reaction temperature is 4°C.

[0041] (4) Use sodium borohydride (NaBH 4 ) reduction, PBS dialyzed overnight.

[0042] 2. Pacific Green is a bright green fluorescent dye, which is excited by the ultraviolet laser 405nm line, and the excitation / emission wavelength is 411 / 510nm. Conjugates of this dye are strongly fluorescent...

Embodiment 2

[0071] 1. Monoclonal antibody-labeled dextran amplification process:

[0072] (1) Dissolve dextran (DeX) in 20mM phosphate buffered saline (PBS) with a pH value of 7.0, add sodium periodate (NaIO 4 ) was oxidized, and the reaction was stirred at 30°C to 37°C in the dark for 2-3 hours and then terminated by adding excess ethylene glycol; the same PBS as the DeX matrix was used for dialysis overnight.

[0073] (2) Dialyze the required antibody against PBS overnight.

[0074] (3) After the oxidized DeX and the antibody were mixed and reacted, an appropriate amount of sodium cyanoborohydride (NaCNBH 3 ) The reaction time in the dark is 1.5-3 hours, and the reaction temperature is 4°C.

[0075] (4) Use sodium borohydride (NaBH 4 ) reduction, PBS dialyzed overnight.

[0076] 2. Amplified antibody-labeled fluorescein:

[0077] (1) Ki67 antibody labeled Pacific Green

[0078] ①Preparation of 1M sodium bicarbonate solution: Weigh about 84mg of sodium bicarbonate, add 1ml of deion...

Embodiment 3

[0101] 1. Monoclonal antibody-labeled dextran amplification process:

[0102] (1) Dissolve dextran (DeX) in 20mM phosphate buffered saline (PBS) with a pH value of 7.0, add sodium periodate (NaIO 4 ) was oxidized, and the reaction was stirred at 30°C to 37°C in the dark for 2-3 hours and then terminated by adding excess ethylene glycol; the same PBS as the DeX matrix was used for dialysis overnight.

[0103] (2) Dialyze the required antibody against PBS overnight.

[0104] (3) After the oxidized DeX and the antibody were mixed and reacted, an appropriate amount of sodium cyanoborohydride (NaCNBH 3 ) The reaction time in the dark is 1.5-3 hours, and the reaction temperature is 4°C.

[0105] (4) Use sodium borohydride (NaBH 4 ) reduction, PBS dialyzed overnight.

[0106] 2. Amplified antibody-labeled fluorescein:

[0107] (1) Ki67 antibody labeled Pacific Green

[0108] ①Preparation of 1M sodium bicarbonate solution: Weigh about 84mg of sodium bicarbonate, add 1ml of deion...

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Abstract

The present invention provides an immunofluorescent double staining kit for cervical cancer auxiliary diagnosis. The kit comprises a mixed primary antibody working solution consisting of a glucan labeled Ki67 monoclonal antibody and a glucan labeled P16INK4A monoclonal antibody, and a DAPI staining solution. The kit simultaneously and efficiently excites multi-color fluorescent light, and all colors can be showed by only one kind of excitation light, so that wavebands do not need to be switched and an ordinary microscope is directly used for observation and shooting; a secondary antibody incubation process is removed, thereby shortening the operation time of the whole process; and the operation is simple, and the sensitivity and the accuracy are high.

Description

technical field [0001] The invention belongs to the technical field of liquid-based cytology immunofluorescence staining, and relates to an immunofluorescence double-staining kit for auxiliary diagnosis of cervical cancer. Background technique [0002] Cervical cancer is the most common gynecological malignancy, and its incidence is second only to breast cancer, seriously endangering women's health. The occurrence and development of cervical cancer is a long and gradual process, and it takes about ten years to develop from precancerous lesions to cancer. However, cervical cancer is the only clear cause, and it is expected to become the first tumor to be conquered by humans. Therefore, prevention and early screening are of great significance to reduce the incidence and mortality of cervical cancer. [0003] At present, the main methods for the diagnosis of cervical cancer are: cervical cytology examination, human papillomavirus (HPV) detection, colposcopy, pathological biop...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/574G01N33/577G01N33/533
CPCG01N33/57411G01N33/577G01N33/533
Inventor 徐丽红李文成孙康俊黄宝福
Owner 江苏美克医学技术有限公司
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