In vitro micronucleus assay for the analysis of total particulate matterin cigarette smoke: comparison of flow cytometry and laser scanningcytometry with microscopy

Yao, Jianhua, Gao, Qian, Mi, Qili, Li, Xuemei, Miao, Mingming, Cheng, Peng and Luo, Ying (2013) In vitro micronucleus assay for the analysis of total particulate matterin cigarette smoke: comparison of flow cytometry and laser scanningcytometry with microscopy. Mutation Research/Genetic Toxicology and Environmental Mutagenesis, 755 2: 120-125. doi:10.1016/j.mrgentox.2013.05.017


Author Yao, Jianhua
Gao, Qian
Mi, Qili
Li, Xuemei
Miao, Mingming
Cheng, Peng
Luo, Ying
Title In vitro micronucleus assay for the analysis of total particulate matterin cigarette smoke: comparison of flow cytometry and laser scanningcytometry with microscopy
Formatted title
In vitro micronucleus assay for the analysis of total particulate matterin cigarette smoke: comparison of flow cytometry and laser scanningcytometry with microscopy
Journal name Mutation Research/Genetic Toxicology and Environmental Mutagenesis   Check publisher's open access policy
ISSN 1383-5718
Publication date 2013-08-15
Year available 2013
Sub-type Article (original research)
DOI 10.1016/j.mrgentox.2013.05.017
Volume 755
Issue 2
Start page 120
End page 125
Total pages 6
Place of publication Amsterdam, Netherlands
Publisher Elsevier
Collection year 2014
Language eng
Formatted abstract
The possible genotoxicity of the total particulate matter (TPM) in cigarette smoke has typically been evaluated using the in vitro micronucleus assay. In recent years, automated scoring techniques have been developed to replace the manual counting process in this assay. However, these automated scoring techniques have not been applied in routine genotoxicity assays for the analysis of TPM to improve the assay efficiency. Chinese hamster ovary (CHO) cells were treated with TPM produced from 14 types of cigarettes at five concentrations (25–200 μg/ml) without exogenous metabolic activation. The three following methods were used to score the micronucleus (MN) frequency: (a) flow cytometry with SYTOX and EMA dyes, which differentially stain micronuclei and apoptotic/necrotic chromatin to enhance assay reliability; (b) laser scanning cytometry with FITC and PI dyes, which is a system that combines the analytical capabilities of flow and image cytometry; and (c) visual microcopy with Giemsa dye. The test results obtained using the three methods were compared using correlation analysis. The key findings for this set of compounds include the following: (a) both flow cytometry- and laser scanning cytometry-based methods were effective for MN identification, (b) the three scoring methods could detect dose-dependent micronucleus formation for the 14 types of TPM, and (c) the MN frequencies that were measured in the same samples by flow cytometry, laser scanning cytometry, and visual microscopy were highly correlated, and there were no significant differences (p > 0.05). In conclusion, both flow cytometry and laser scanning cytometry can be used to evaluate the MN frequency induced by TPM without exogenous metabolic activation. The simpler and faster processing and the high correlation of the results make these two automatic methods appropriate tools for use in in vitro micronucleus assays for the analysis of TPM using CHO cells.
Keyword Flow cytometry
Laser scanning cytometry
Cigarette smoke
Total particulate matter
Micronucleus
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Official 2014 Collection
National Research Centre for Environmental Toxicology Publications
 
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