Using fluorescence for improvement of the quantitative analysis of micronucleus in cell culture.
The micronucleus test is of unquestionable importance, being widely used in studies with several purposes. Standardization of the technique has been proposed by several groups mainly when in vivo assays are performed. In cell cultures the staining method that predominates is the Feulgen's reaction, since it is specific for DNA. In this work, we evaluated the use of two fluorescent stains, SYTOX green and propidium iodide, in substitution for the Feulgen's reaction. Based on the results we concluded that the fluorescence microscopy allows the reliable detection of micronuclei, presenting the following advantages: the technique is easy and fast to perform, it avoids acid treatments; the fluorescence is long lasting; micronucleus counting becomes easier and more reliable by the use of RNAase treatment; the technique is more sensitive to detect the smallest micronuclei.
Dias
Viviane Miranda
VM
Department of Cellular and Development Biology, Institute of Biomedical Sciences, University of São Paulo (USP), Av. Prof. Lineu Prestes, 1524, São Paulo-CEP: 05508-900, SP, Brazil. vi_dias@yahoo.com.br
Manelli-Oliveira
Renata
R
Machado-Santelli
Gláucia Maria
GM
D000818:Animals; D016590:Aphidicolin / Q000378:metabolism; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D004791:Enzyme Inhibitors / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism*; D006801:Humans; D007202:Indicators and Reagents / Q000378:metabolism*; D015162:Micronucleus Tests / Q000379:methods*; D008856:Microscopy, Fluorescence / Q000379:methods*; D009930:Organic Chemicals; D011419:Propidium / Q000378:metabolism*; D012394:Rosaniline Dyes / Q000378:metabolism
D004791:Enzyme Inhibitors; C007722:Feulgen stain; D005456:Fluorescent Dyes; D007202:Indicators and Reagents; D009930:Organic Chemicals; D012394:Rosaniline Dyes; C402795:SYTOX Green; D011419:Propidium; D016590:Aphidicolin