Masayuki Nagahashi1, Yoshifumi Shimada2, Hiroshi Ichikawa2, Satoru Nakagawa3, Nobuaki Sato4, Koji Kaneko4, Keiichi Homma5, Takashi Kawasaki5, Keisuke Kodama6, Stephen Lyle7, Kazuaki Takabe8, Toshifumi Wakai2. 1. Division of Digestive and General Surgery, Niigata University Graduate School of Medical and Dental Sciences, Niigata City, Niigata, Japan. Electronic address: mnagahashi@med.niigata-u.ac.jp. 2. Division of Digestive and General Surgery, Niigata University Graduate School of Medical and Dental Sciences, Niigata City, Niigata, Japan. 3. Department of Surgery, Niigata Cancer Center Hospital, Niigata City, Niigata, Japan. 4. Department of Breast Oncology, Niigata Cancer Center Hospital, Niigata City, Niigata, Japan. 5. Department of Pathology, Niigata Cancer Center Hospital, Niigata City, Niigata, Japan. 6. Diagnostics Research Department, Life innovation Research Institute, Denka innovation Center, Denka Co., Ltd, Machida City, Tokyo, Japan. 7. Department of Molecular, Cell and Cancer Biology, University of Massachusetts Medical School, Boston, Massachusetts; KEW, Inc, Boston, Massachusetts. 8. Breast Surgery, Department of Surgical Oncology, Roswell Park Cancer Institute, Buffalo, New York; Department of Surgery, University at Buffalo Jacobs School of Medicine and Biomedical Sciences, The State University of New York, Buffalo, New York.
Abstract
INTRODUCTION: Precision medicine is only possible in oncology practice if targetable genes in fragmented DNA, such as DNA from formalin-fixed paraffin-embedded (FFPE) samples, can be sequenced using next generation sequencing (NGS). The aim of this study was to examine the quality and quantity of DNA from FFPE cancerous tissue samples from surgically resected and biopsy specimens. METHODS: DNA was extracted from unstained FFPE tissue sections prepared from surgically resected specimens of breast, colorectal and gastric cancer, and biopsy specimens of breast cancer. A total quantity of DNA ≥60 ng from a sample was considered adequate for NGS. The DNA quality was assessed by Q-ratios, with a Q-ratio >0.1 considered sufficient for NGS. RESULTS: The Q-ratio for DNA from FFPE tissue processed with neutral-buffered formalin was significantly better than that processed with unbuffered formalin. All Q-ratios for DNA from breast, colorectal and gastric cancer samples indicated DNA levels sufficient for NGS. DNA extracted from gastric cancer FFPE samples prepared within the last 7 years is suitable for NGS analysis, whereas those older than 7 years may not be suitable. Our data suggested that adequate amounts of DNA can be extracted from FFPE samples, not only of surgically resected tissue but also of biopsy specimens. CONCLUSIONS: The type of formalin used for fixation and the time since FFPE sample preparation affect DNA quality. Sufficient amounts of DNA can be extracted from FFPE samples of both surgically resected and biopsy tissue, thus expanding the potential diagnostic uses of NGS in a clinical setting.
INTRODUCTION: Precision medicine is only possible in oncology practice if targetable genes in fragmented DNA, such as DNA from formalin-fixed paraffin-embedded (FFPE) samples, can be sequenced using next generation sequencing (NGS). The aim of this study was to examine the quality and quantity of DNA from FFPE cancerous tissue samples from surgically resected and biopsy specimens. METHODS: DNA was extracted from unstained FFPE tissue sections prepared from surgically resected specimens of breast, colorectal and gastric cancer, and biopsy specimens of breast cancer. A total quantity of DNA ≥60 ng from a sample was considered adequate for NGS. The DNA quality was assessed by Q-ratios, with a Q-ratio >0.1 considered sufficient for NGS. RESULTS: The Q-ratio for DNA from FFPE tissue processed with neutral-buffered formalin was significantly better than that processed with unbuffered formalin. All Q-ratios for DNA from breast, colorectal and gastric cancer samples indicated DNA levels sufficient for NGS. DNA extracted from gastric cancer FFPE samples prepared within the last 7 years is suitable for NGS analysis, whereas those older than 7 years may not be suitable. Our data suggested that adequate amounts of DNA can be extracted from FFPE samples, not only of surgically resected tissue but also of biopsy specimens. CONCLUSIONS: The type of formalin used for fixation and the time since FFPE sample preparation affect DNA quality. Sufficient amounts of DNA can be extracted from FFPE samples of both surgically resected and biopsy tissue, thus expanding the potential diagnostic uses of NGS in a clinical setting.
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