Method for preservation of DNA stability of liquid-based cytology specimens from a lung adenocarcinoma cell line.

Abstract

Liquid-based cytology (LBC) specimens of lung adenocarcinoma have the potential to be widely used for genetic analysis. However, formaldehyde contained in some LBC preservation solutions can cause DNA fragmentation during specimen storage, rendering the samples unsuitable for molecular analysis. To investigate a novel preservation technique for improved DNA stability, which was evaluated by mutation analysis of epidermal growth factor receptor (EGFR) gene in human lung adenocarcinoma cell lines. Cells were fixed in CytoRich Red preservation solution. After 30min of fixation, cells were either stored using the conventional method (suspended in preservation solution) or washed in phosphate-buffered saline and stored as a cell pellet (newly proposed method). The effect of storage was evaluated after 5, 7, and 9days of storage at ambient temperature. The cell pellet group was also tested after 14 and 28days. Specifically, we evaluated the DNA stability, DNA yield, and sample suitability for polymerase chain reaction (PCR), and EGFR mutation detection. The DNA yields and degree of stability from the cell pellet group were higher than those from the suspension group at every time point examined. PCR amplification from the cell pellet group was successful up to day 28. Mutation detection using the Cycleave PCR method indicated that the Ct values of the cell pellet group were significantly lower than those of the suspension group. Storing LBC specimens as a cell pellet post-fixation can maintain the DNA quality for a longer period than the conventional method, making it a promising strategy for molecular analysis.