| Popis: |
Background: There are enormous formalin fixed paraffin embedded tissue archives and constantly growing number of methods for molecular analyses but, isolation of DNA from this tissue is still challenging due to the damage effect of formalin on DNA. To determine the extent to which DNA purity, quantity and integrity depends on the process of fixation in formalin, and to what extent on the process of tissue paraffin embedding, we compared the quality of DNA isolated from fixed tissues and DNA isolated from tissues embedded in paraffin blocks after fixation. Methods and Results: Heart, liver and brain tissues obtained from healthy people who suddenly died a violent death were fixed in 10% buffered formalin as well as in 4% unbuffered formalin 6h, 1-7 days (every 24h), 10, 14, 28 days and 2 months. Also the same tissues were fixed in 4% unbuffered formalin and embedded in paraffin block and stored from few months to 30 years. Yield and purity of the DNA samples isolated from these tissues were measured using spectrophotomer The PCR amplification of the hTERT gene was performed to evaluate the degree of DNA molecule fragmentation. Although the purity of the DNA isolated from almost all tissue samples is satisfactory, the DNA yields changes significantly. Conclusion: The largest decrease in DNA yield was observed after tissue fixation in formalin, especially with prolonged formalin fixation, and additionally after paraffin embedding of tissue. DNA integrity also depends on time of tissue formalin fixation and the age of paraffin blocks. |
