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Cancer promotion activity quantitative determination method and screening method of cancer promoter

A quantitative detection and activity technology, applied in the field of biomedicine, can solve the problems of prolonged detection time, time-consuming and laborious, unable to meet the actual detection of fast and large samples, etc., and achieve the effect of quantitative detection.

Inactive Publication Date: 2015-04-15
PEKING UNIV
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  • Application Information

AI Technical Summary

Problems solved by technology

However, the microinjection method requires a microscopic operating system, which requires high precision in operation; the scratch dye labeling and tracing method has high requirements for cell scratches, and it takes time and effort for many samples.
[0016] Although the above methods are complementary to each other, they all have their own shortcomings. Due to the limitation of the measurement period, detection system, equipment and / or operator skills, the detection of the function of the intercellular plasma membrane channel in each method requires specific Samples are analyzed one by one, and the detection time is greatly extended, which cannot meet the needs of rapid and large-scale actual detection. Therefore, it is necessary to develop fast, high-throughput quantitative detection methods for the function of intercellular membrane channels and cancer-promoting agent screening methods. It is of great value for the practical application of evaluating the cancer-promoting activity of environmental chemicals

Method used

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  • Cancer promotion activity quantitative determination method and screening method of cancer promoter
  • Cancer promotion activity quantitative determination method and screening method of cancer promoter
  • Cancer promotion activity quantitative determination method and screening method of cancer promoter

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Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0062] 1. Establish a stable fluorescent probe-labeled cell model

[0063] 1.1 Cell culture

[0064] Select human immortalized epidermal HaCat cells (CCTCC, China Type Culture Collection Center), use MEM medium containing 10% fetal bovine serum and penicillin-streptomycin double antibody (penicillin 100U / mL; streptomycin 100 μg / mL), At 37°C, 5% CO 2 cultivated under conditions. The cells in the logarithmic growth phase were digested with 0.25% trypsin (containing 0.03% EDTA), and passaged once every 2-3 days.

[0065] 1.2 Fluorescent probe preparation

[0066] Calcein AM (C-3099, Molecular Probes) solution (1 mM) was aliquoted and stored at -20°C in the dark for future use. DiI (D-282, Molecular Probes) powder was prepared into 2mM stock solution with DMSO, and stored at -20°C in the dark for future use after aliquoting.

[0067] Probe diluent: Glucose was dissolved in sterilized double distilled water to prepare a 0.3M glucose use solution, and stored at 4°C for later us...

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Abstract

A quantitative method for detection of cancer promotion activity of analytes. The method comprises the following steps: selecting a suitable cell model, and dividing the cells into donor cells and recipient cells according to the demand; adding two fluorescent probes in the donor cells and incubating; conducting mixed culture on the donor cells and recipient cells according to the proportion; adding the analyte into a test group of mixed culture cells, wherein the control group is not supplemented; incubating for a certain time, and measuring the number of one fluorescent probe, in the donor cells of test group and control group, transferring to the single positive cells in recipient cells by using a flow cytometry; calculating Te (Transfer effects) values of the two groups, and respectively representing with Tecon and Tetest, wherein Te equals to the number of single positive cells / number of single donor cell * 100%; using TeR to represent the relative value of Te of the analyte, wherein TeR equals Tetest / Tecon * 100%; and evaluating the tumor promotion activity of the analyte according to TeR of the analyte, wherein lower TeR indicates higher activity of the analyte. The invention also relates to application of the above method to in vitro screening of cancer promoters; and when Te is less than 90%, the analyte can be identified as a cancer promoter.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to a quantitative detection method for cancer-promoting activity and a screening method for cancer-promoting agents. Background technique [0002] Environmental pollution is an issue of great concern in today's society. A large number of studies at home and abroad have shown that more than 70% of carcinogenic and cancer-promoting factors are closely related to environmental pollution and human behavior during the occurrence and development of cancer. Carcinogenic substances pollute the environment is one of the important reasons to increase the incidence of cancer. Environmental pollutants that cause human cancer can pollute the environment through air, water, and soil, and can cause cancer in a wide range of sites, including lung, skin, bladder, larynx, ovary, breast, nasopharynx, kidney, and hematopoietic system. The World Cancer Report 2014 by the International ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64
Inventor 邓芙蓉魏红英郭新彪杨迪
Owner PEKING UNIV
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