Cytochemical Staining of DNA

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BACKGROUND

Felugen reaction or cytochemical staining of DNA is a method of identifying DNA or other chromosomal material in a given cell specimen. It mainly involves the acid hydrolysis of DNA and mostly involves strong acids. It is a semi-quantitative DNA. So, the basic objective of this test is to study in details about the cytochemical staining of DNA.

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REQUIREMENTS

Sample:                   Specimen cell

Chemicals:              Aqueous hydrochloric acid

• Schiff Reagent – Pour 200 ml of boiled distilled water over 1 g basic fuchsin. Shake it properly and cool it to 50^oC, filter it and to it add 30 ml of 1 N HCL. Allow it to cool down to room temperature and add 1 g of potassium metabisulphite. Allow this solution to stand overnight in dark till a light straw or faint pink colour develops.
  
• Xylene
  
• Alcohol, Ethyl denatured 100%
  
• Alcohol Ethyl denatured 95%
  
• Light Green SF yellowish stain 0.2%, Aqueous
  

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PROCEDURE

First make hydrochloric acid working solution and mix it well. For the solution you have to mix 16 ml of hydrochloric acid, 20% aqueous with 24 ml of distilled water. Pre-heat the solution and maintain it at 60^oC before using.¹ Thoroughly deparaffinize the slide sections in three alterations of xylene for about 3 minutes each. Hydrate it through two alterations each in 100% and 95% ethyl alcohol with 10 dips each. Wash it well with distilled water.² Then next hydrolyze each sections in working solution of hydrochloric acid at 60^oC for about 10 minutes. Now place the slides in the Schiff reagent, McManus for about 45 minutes and then wash it with distilled water.³ Counterstain it in light green SF yellowish stain 0.2% aqueous for about 1 minute. Now dehydrate it in two changes each of 95% and 100% ethyl alcohol. Clean in about three alterations of xylene with 10 dips each with the help of compatible mounting medium.⁴

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CONCLUSION

More the staining intensity more is the concentration of the DNA in this Feulgen reaction.

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REFERENCES

1. Bancroft, John D., and Marilyn Gamble. Theory and Practice of Histological Techniques. 6th ed. Oxford: Churchill Livingstone Elsevier, 2008.224-225.
   
2. Carson, Freida L., and Christa Hladik Cappellano. Histotechnology: A Self-Instructional Text. 4th ed. Chicago: ASCP Press, 2015. 126-127.
   
3. Sheehan, Dezna C., and Barbara B. Hrapchak. Theory and Practice of Histotechnology. 2nd ed. St. Louis: Mosby, 1980. 150.
   
4. Modifications developed by Newcomer Supply Laborator.