Method for determining and evaluating cytotoxicity caused by cigarette smoke

A technology of cigarette smoke and cytotoxicity, which is applied in the field of cigarette smoke toxicology research, can solve the problems of individual differences, inconvenient material collection, and long experiment time, and achieve the effects of short action time, shortened color development time, and convenient material collection

Active Publication Date: 2012-03-14
CHINA TOBACCO ZHEJIANG IND
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There are individual differences in the collection of heparin-anticoagulated peripheral blood, which is inconvenient to obtain materials and takes a long time for the experiment.

Method used

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  • Method for determining and evaluating cytotoxicity caused by cigarette smoke
  • Method for determining and evaluating cytotoxicity caused by cigarette smoke
  • Method for determining and evaluating cytotoxicity caused by cigarette smoke

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1 Evaluating the Cytotoxicity of Flue-cured Tobacco Smoke in China

[0024] According to the ISO 3308 suction conditions (each suction volume is 35ml, suction 2s, interval 60s) after continuously smoking 5 cigarettes, take out the Cambridge filter, put it in 10ml DMSO solution, vibrate at 145 rpm for 30 minutes, divide Store in a -70°C freezer for later use.

[0025] The HMy2.CIR human B-lymphoblastoid cell line was purchased from the Cell Bank of the Chinese Academy of Sciences and incubated at 37°C and 5% CO with IMDM medium (containing 10% fetal bovine serum). 2 The cells were cultured in an incubator and subcultured every 2-3 days. Cells in the exponential growth phase were taken for infection, and 5×10 cells were inoculated in each well of a 96-well plate. 4 B lymphocytes, 2.5×10 -3 , 5.0×10 -3 , 7.5×10 -3 , 10.0×10 -3 and 12.5×10 -3 The concentration of cigarettes / ml was tested, each concentration was inoculated into 3 wells, and the exposure time was...

Embodiment 2

[0032] Example 2 Evaluating the Cytotoxicity of Smoke from a Domestic Mixed Type Cigarette

[0033] According to the ISO 3308 suction conditions (each suction volume is 35ml, suction 2s, interval 60s) after continuously smoking 5 cigarettes, take out the Cambridge filter, put it in 10ml DMSO solution, vibrate at 145 rpm for 30 minutes, divide Store in a -70°C freezer for later use.

[0034] The HMy2.CIR human B-lymphoblastoid cell line was purchased from the Cell Bank of the Chinese Academy of Sciences and cultured in IMDM medium (containing 10% fetal bovine serum) at 37°C and 5% CO 2 The cells were cultured in an incubator and subcultured every 2-3 days. Cells in the exponential growth phase were taken for infection, and 5×10 cells were inoculated in each well of a 96-well plate. 4 For B lymphocytes, 2.5×10 -3 , 5.0×10 -3 , 7.5×10 -3 , 10.0×10 -3 and 12.5×10 -3 The concentration of cigarettes / ml was tested, each concentration was inoculated into 3 wells, and the exposu...

Embodiment 3

[0041] Example 3 Evaluating the Cytotoxicity of Flue-cured Tobacco Cigarettes Abroad

[0042] According to the ISO 3308 suction conditions (each suction volume is 35ml, suction 2s, interval 60s) after continuously smoking 5 cigarettes, take out the Cambridge filter, put it in 10ml DMSO solution, vibrate at 145 rpm for 30 minutes, divide Store in a -70°C freezer for later use.

[0043] The HMy2.CIR human B-lymphoblastoid cell line was purchased from the Cell Bank of the Chinese Academy of Sciences and incubated at 37°C and 5% CO with IMDM medium (containing 10% fetal bovine serum). 2 The cells were cultured in an incubator and subcultured every 2-3 days. Cells in the exponential growth phase were taken for infection, and 5×10 cells were inoculated in each well of a 96-well plate. 4 For B lymphocytes, 2.5×10 -3 , 5.0×10 -3 , 7.5×10 -3 , 10.0×10 -3 and 12.5×10 -3 The concentration of cigarettes / ml was tested, each concentration was inoculated into 3 wells, and the exposure...

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Abstract

The invention relates to the field of cigarette smoke toxicology researche, in particular to a method for determining and evaluating cytotoxicity caused by cigarette smoke. The method for determining and evaluating cytotoxicity caused by cigarette smoke comprises the following steps: (1) preparing an extracting solution of cigarette smoke particulate matters; (2) culturing an HMy2.CIR human B lymph mother cell line; (3) contaminating; (4) developing; (5) calculating the cell survival rate; and (6) drawing a graph to calculate IC50. The invention has the advantages that no individual difference exists, thus influences on experiments of the individual difference can be eliminated; materials are convenient to obtain, and the quantity of cells is not limited so that the requirement on a largesample quantity can be met; ethical issues are not involved; the action time with CCK-8 is short, the developing time can be shortened, experimental errors caused by long developing time can be reduced, and an experimental process is quicker.

Description

technical field [0001] The invention relates to the field of cigarette smoke toxicology research, in particular to a method for measuring and evaluating the cytotoxicity caused by cigarette smoke. Background technique [0002] Cytotoxicity plays an important role in understanding the mechanism of action of chemicals on cells and / or tissues, which can affect several pathological processes including carcinogenesis and inflammation. Cytotoxicity can also mediate the effects of other substances such as free radicals, stimulants and genotoxic substances (Bombick, DW et al., chemical and biological studies of a new cigarette that primarily heats tobacco, Food and Chemical Toxicology, 1998, 36, 191-197 .). Cigarette smoke is a complex mixture containing more than 6,000 chemicals, many of which are cytotoxic. Cytotoxicity tests can screen these chemicals to discover potentially harmful toxic effects and provide timely information for the study of the mechanism of chemical toxicity...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/02G01N21/31
Inventor 储国海周国俊楼建林蒋健黄芳芳何继亮
Owner CHINA TOBACCO ZHEJIANG IND
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