Description
This detailed academic note on the Demonstration of Nucleic Acids provides an essential guide to histochemical staining techniques for tertiary students in Histotechnology, Biomedical Science, and Molecular Biology. Derived from the authoritative text "Bancroft's Theory & Practice of Histological Techniques" by Dr. S.K. Suvarna et al., the document systematically outlines methods for identifying and differentiating DNA and RNA in tissue sections and laboratory preparations. It begins by explaining the strong basophilic nature of nucleic acids and the need for specific techniques beyond general basic dyes. The note covers the classic Feulgen technique, which is specific for DNA, relying on acid hydrolysis to expose aldehydes that react with Schiff's reagent. It details the Methyl Green-Pyronin Y method for simultaneously staining DNA green and RNA red, based on the differential polymerization of the two nucleic acids. The Gallocyanin-Chrome Alum method for staining both nucleic acids blue is also explained. Furthermore, the material explores modern fluorescent stains, including Acridine Orange, which distinguishes between double-stranded and single-stranded nucleic acids by emitting green or orange-red fluorescence, respectively, and Ethidium Bromide, the standard for visualizing DNA in agarose gels under UV light. A summary table compares the key stains, their targets, and resulting colours. This resource is vital for diploma and degree students and research scientists to accurately localize and analyze nucleic acids for diagnostic and research purposes. Download these comprehensive notes to master the principles and applications of nucleic acid histochemistry.
What's inside
Demonstration of Nucleic Acids Introduction • Nucleic acids are molecules that store information for cellular growth and reproduction. • Two nucleic acids are: 1. DNA 2. RNA • They consist of : sugar (Deoxyribose/Ribose), phosphate and nitrogenous base. Staining Techniques for NAs • NAs are strongly basophilic and will stain with almost any basic dye. • This is fine for morphological purposes but basophilia is not limited to NAs and needs controlling in order to get specific identification. Feulgen Technique Purpose • Used for the identification of DNA in paraffin-embedded tissue or cell specimens. • The method is specific for DNA and will not stain RNA. Feulgen Technique Principle of the Feulgen Method • HCl hydrolyze the deoxyribose sugar of DNA into an aldehyde. • Results in the cleavage of the nitrogen bases. • Aldehyde reacts with Schiff’s reagent, which specifically binds to aldehydes. Feulgen Technique • RNA is not hydrolysed by the HCl treatment, and, thus the reaction is DNA specific • A light green counterstain is often applied to the tissue section to achieve a better contrast. Feulgen Technique • Is a semi-quantitative technique • If the only aldehydes remaining in the cell are produced from hydrolysis of DNA • DNA in each cell can be measured by microdensitometry. Results DNA………Red-Purple Cytoplasm…..Green Methyl Green-Pyronin Technique Purpose • For demonstration of both DNA and RNA Principle • The method uses two different basic dyes to stain the different NAs and distinguish DNA and RNA. • Methyl green (a triaryl methane dye) = DNA • Pyronin Y(a xanthene dye) = RNA • The reason that two basic dyes bind preferentially is suggested to be related to the different degrees of polymerization observed in DNA and RNA. Methyl Green-Pyronin Technique When used at pH 4.6 the dyes compete for binding to the NAs DNA → green colour RNA → red colour Methyl Green-Pyronin Technique
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