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Method for testing in-vitro cytotoxicity of mouth cigarette extract

A cytotoxicity and testing method technology, applied in the field of in vitro toxicology evaluation of snus, can solve the problem of no in vitro toxicity testing method, etc., and achieve the effects of good repeatability of test results, strong stability and simple experimental process

Inactive Publication Date: 2016-12-07
ZHENGZHOU TOBACCO RES INST OF CNTC
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Problems solved by technology

Reports on the health effects of snus are mainly based on epidemiological data and the results of laboratory animal experiments on a single compound, while there are few literature reports on the in vitro toxicity test of snus, and there is no uniform standard in vitro toxicity test method

Method used

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  • Method for testing in-vitro cytotoxicity of mouth cigarette extract
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  • Method for testing in-vitro cytotoxicity of mouth cigarette extract

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Embodiment Construction

[0032] The present invention will be further described below in conjunction with embodiment accompanying drawing:

[0033] Specific implementation one

[0034] Deionized water and dimethyl sulfoxide (DMSO) were selected to extract a commercially available snus sample to obtain water-soluble extracts and DMSO extracts, respectively. CCK-8 assay was used to detect the effect of the extract on the survival rate of Chinese hamster ovary cells (CHO cells). CHO cells at 1 x 10 4 cells / well were seeded in a 96-well plate, at 37°C, 5% CO 2 After culturing for 24 h under the same conditions, the cells were infected for 24 h. The exposure doses of water-soluble extracts were: 0 (negative control), 2500, 5000, 10000, 20000 and 40000 μg / mL; the exposure doses of DMSO extracts were: 0 (negative control), 250, 500, 1000 , 2000, and 4000 μg / mL. After 24 hours of exposure, 10 μL of CCK-8 reagent was added to each well, and the cells were incubated at 37 °C, 5% CO 2 Incubate for about 3 ...

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Abstract

A method for testing in-vitro cytotoxicity of mouth cigarette extract comprises the following steps that 1, the mouth cigarette extract is prepared; 2, cell inoculation culture is carried out; 3, the mouth cigarette extract is subjected to contamination; 4, a CCK-8 dyeing experiment is carried out; 5, a result is obtained, and analysis is carried out. The method is characterized in that in-vitro cytotoxicity testing of mouth cigarettes is established for the first time, when testing is carried out, the step of washing cells many times is omitted, and the experiment process is simpler and more convenient. A CCK-8 reagent can be directly added into a cell culture medium, no liquid replacement step is needed, the generated water-soluble formazan product can be directly subjected to light absorption value detection, a dissolving step before detection is omitted, and the experiment period and time are shortened; continuous light absorption value detection can be carried out many times, no additional experiment step needs to be added, and convenience is provided for looking for a proper detecting time point; the toxicity of the CCK-8 reagent on the cells is small, the influence of the background and experiment errors are avoided, and the testing result is good in repeatability and high in stability and sensitivity.

Description

technical field [0001] The invention relates to the research field of in vitro toxicological evaluation of snus, in particular to an in vitro cytotoxicity testing method of snus extract. Background technique [0002] Snus is a new type of tobacco product that consumes tobacco through non-combustion routes. It does not require combustion, has significantly fewer harmful components, and does not produce "environmental tobacco smoke". The consumption of snus is a tradition in many countries. However, research reports have shown that snus can also cause adverse effects on human health, can cause cancer in the oral cavity, digestive tract, and other parts of the human body, can be addictive, and may be a risk factor for cardiovascular and cerebrovascular diseases. In fact, there are many types of snus, and the composition and carcinogenic ability of different types vary greatly. According to available data, there are 28 known carcinogens or potential carcinogens in snus, inclu...

Claims

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Application Information

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IPC IPC(8): C12Q1/02
CPCG01N33/5014
Inventor 李翔赵俊伟尚平平乔梁峻华辰风谢复炜刘惠民
Owner ZHENGZHOU TOBACCO RES INST OF CNTC
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