PCR amplification from paraffin-embedded tissues: recommendations on fixatives for long-term storage and prospective...

Area of Science:

• Molecular Biology
• Histology
• Genetics

Background:

• Polymerase chain reaction (PCR) enables molecular studies on archival tissues.
• DNA damage from certain fixation methods can hinder PCR analysis.
• Optimizing fixation and storage is crucial for reliable molecular analysis of historical samples.

Purpose of the Study:

• To evaluate the impact of various fixatives and storage durations on DNA amplifiability from paraffin-embedded tissues.
• To determine optimal fixation methods for preserving DNA integrity for PCR analysis.
• To provide guidance for molecular studies using archival specimens.

Main Methods:

• Tissue samples were fixed for 2 hours to 30 days using 10% buffered-neutral formalin (BNF), 95% ethanol, acetone, and OmniFix.
• Paraffin embedding was performed after fixation.
• DNA amplification was assessed using PCR for fragments ranging from 268 to 1327 bp.

Main Results:

• Formalin fixation progressively reduced DNA amplifiability, with only 268 bp fragments amplifiable after 30 days.
• OmniFix and acetone allowed amplification of 989 bp fragments after 30 days.
• 95% ethanol fixation efficiently yielded DNA amplification products up to 1327 bp, even after 30 days.

Conclusions:

• 95% ethanol is a superior fixative for maintaining DNA integrity for long-term PCR analysis of archival tissues.
• Ethanol fixation is particularly valuable for remote locations lacking refrigeration or immediate processing capabilities.
• The study offers critical data for selecting appropriate fixation techniques in molecular pathology and historical sample research.