Primary cell Feulgen reaction shows DNA
Fundamental:
The traditional method of displaying DNA is the Feulgen reaction. The section is first treated with dilute hydrochloric acid. After the DNA is hydrolyzed by a weak acid (1 mol/L HCL), the DNA molecule is linked to deoxyribose and ruthenium at 60 °C. Upon opening, one end of the deoxyribose releases an aldehyde group and reacts in situ with the Schiff reagent to form a purplish red product. The principle reacts with PAS to make the nuclear DNA appear purple-red. RNA is not affected during this process, so staining is DNA specific. Accurate temperature and hydrochloric acid concentration, the appropriate hydrolysis time is the key to success. Excessive or insufficient hydrolysis reduces the intensity of staining.
Formulation reagents:
1. 1mol/L HCL: 8.5ml concentrated hydrochloric acid and 91.5ml distilled water are mixed;
2. 0.5% bright green aqueous solution;
Dyeing steps:
1. Take adherent primary cells cultured on coverslips and rinse three times with Hanks solution;
2. Fix in Carnoy fixative for 10 min, then wash three times with distilled water;
3. Move into a 60 ° C concentration of 1 mol / L HCL and let stand for 10 min;
4. Rinse several times with distilled water;
5. Place in Schiff reagent for 30-60 minutes, check the coloration at any time;
6. Rinse with tap water for 5 min;
7. Rinse with distilled water;
8. Dehydration of different concentration gradient ethanol solutions;
9. Use xylene transparent, glued to seal the sheet;
result:
The DNA in the primary nucleus is purple-red and the cytoplasm is green;
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