Good DNA- and sufficient RNA integrity in formalin-free paraffin-embedded tissues.

Abstract

In recent decades, the field of pathology has been enriched by the development of molecular biology, enabling increased specificity in cancer diagnosis and personalized therapy based on the molecular profile of tumor cells. Traditional tissue embedding techniques, routinely using toxic and even carcinogenic substances, in particular formaldehyde, are still standard practice for the field of pathology and consequently for molecular pathology. Formaldehyde damages DNA and RNA, resulting in suboptimal molecular diagnostics, possibly impacting cancer diagnosis and treatment quality. A novel tissue embedding technique based on supercritical CO2 can be used in a formaldehyde-free manner, producing high-quality samples suitable for basic pathology techniques. Early data indicates that the DNA isolated from tissues processed in this manner can be used for molecular pathology, but detailed DNA analysis lacked, and RNA had not been analyzed at all. Here we performed integrity analysis of both the DNA and RNA isolated from tissues 7 years, 2 years, and 3 days after embedding and compared non-fixed and formalin-fixed tissues. The results show that this novel embedding technique, used in xylene and formalin-free manner, results in DNA of higher quality than that of conventionally processed tissues, as shown by higher DIN values. On the other hand, RNA quality, as shown by RIN values, was generally lower in 3-day and 2-year old, non-fixed (NFPE) tissues compared to formalin-fixed (FFPE) tissues but similar in 7 year-old tissues. RNA isolated from 3-day, 2-year, and even most 7-year-old non-formalin fixed tissues was still suitable for the main downstream molecular technique Next-Generation-Sequencing.