Document Type : Letters to the Editor
Author
Dept. of Oral and Maxillofacial Pathology, Faculty of Dentistry, Shahed University, Tehran, Iran
Abstract
Among different staining methods used to demonstrate the nuclear abnormalities, Feulgen is one of the most reliable method. Feulgen staining is specific, sensitive method for evaluating the DNA damages.It has been shown that using non-DNA specific stains for monitoring the nuclear anomalies lead to false-positive or false-negative results. From self-experience, immersing the stained slides in hematoxylin for 3-5 minutes in final step after staining the slides with light green, provides a greater ground contrast and very better visualizing of nucleus. By this method, a clear distinction can be made between micronuclei and other structures suggestive of a micronucleus.
Highlights
1- This idea is original.
2- It can be a modified method for better visualizing of nucleus.
3- By this method, a clear distinction can be made between micronuclei and other structures suggestive of a micronucleus.
Keywords
Dear Editor in Chief
Micronuclei (MN) are fragments of chromosomes that fail to encompass into the nucleus and remains in the cytoplasm in cell division course (1). Monitoring MN in oral exfoliated epithelial cells for detecting exposed individuals to genotoxic agents was proposed by Stich et al. in 1982 for the first time (2).
The genotoxic agents of tobacco, waterpipe and even environmental contaminants are recognized as initiator factors in producing the aberrations that results in MN production (3-5).Assessing the frequency of MN in oral exfoliated cells is a non-invasive reliable indicator of genotoxic damage in human (6).
Among different staining methods to demonstrate the nuclear abnormalities, Feulgen is one of the most reliable method. The Feulgen
staining is a specific and sensitive method for evaluating DNA damages. It has been shown that using non-DNA specific stains to monitor nuclear anomalies lead to false-positive or false-negative results (7).
In a routine Feulgen staining technique, slides are immersed in 5 mol/L HCl for 15 minutes, rinsed with distilled water for 3 minutes, stained with Schiff's reagent for 90 minutes, washed for 10 minutes and finally stained with 1% light green for 15 minutes (7-8). Using the Feulgen staining, nucleus and micronuclei appear magenta and cytoplasm seems green.
From self-experience, immersing the stained slides in hematoxylin for 3-5 minutes and eosin for few seconds in the final step after staining the slides with light green, provides a greater ground contrast and very better visualizing of nucleus (Fig1).
Using this method, the shape and outline of nucleus appear very clear and precise. Nuclear changes as karyorrhexis, karyolysis and pyknosis appear very characteristic and distinctive. By this method, a clear distinction can be made between micronuclei and other structures suggestive of a micronucleus. This experience of using hematoxylin-eosin stain in routine Feulgen staining protocol can be a modified method for Feulgen staining technique for better visualization of nucleus.
- Cerqueira EM, Santoro CL, Donozo NF, Freitas BA, Pereira CA, Bevilacqua RG, etal. Genetic damage in exfoliated cells of the uterine cervix. Association and interaction between cigarette smoking and progression to malignant transformation? Acta Cytol 1998;42 (3):639-649.
- Stich HF, Curtis JR, Parida BB. Application of the micronucleus test to exfoliated cells of high cancer risk groups: tobacco chewers. Int J Cancer 1982; 30 (5):553-559.
- Naderi NJ, Farhadi S, Sarshar S. Micronucleus assay of buccal mucosa cells in smokers with the history of smoking less and more than 10 years. Indian J Pathol Microbiol 2012; 55(4):433-438.
- El-Setouhy M, Loffredo CA, Radwan G, Abdel Rahman R, Mahfouz E, Israel E, etal. Genotoxic effects of waterpipe smoking on the buccal mucosa cells. Mutat Res 2008; 655(1-2):36-40.
- Salama SA, Serrana M, Au WW. Biomonitoring using accessible human cells for exposure and health risk assessment. Mutat Res 1999; 436(1):99-112.
- Mr P, Guruprasad Y, Jose M, Saxena K, Deepa K, Prabhu V.Comparative study of genotoxicity in different tobacco related habits using micronucleus assay in exfoliated buccal epithelial cells. J Clin Diagn Res 2014; 8(5):ZC21-24
- Nersesyan A, Kundi M, Atefie K, Schulte-Hermann R, Knasmüller S. Effect of staining procedures on the results of micronucleus assays with exfoliated oral mucosa cells. Cancer Epidemiol Biomarkers Prev 2006; 15: 1835-1840.
- Thomas P, Hecker J, Faunt J, Fenech M. Buccal micronucleus cytome biomarkers may be associated with Alzheimer's disease. Mutagenesis 2007; 22(6):371-379.
)