期刊
CLINICA CHIMICA ACTA
卷 488, 期 -, 页码 129-134出版社
ELSEVIER SCIENCE BV
DOI: 10.1016/j.cca.2018.11.002
关键词
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资金
- Yamanashi Prefecture
- Japan Society for the Promotion of Science (JSPS) KAKENHI Early-Career Scientists [JP18K16292]
- YASUDA Medical Foundation
- Uehara Memorial Foundation
- Research Grant for Young Scholars
Background: Tumor genetic alterations are determined to aid in selecting therapy and predicting prognosis. In routine clinical practice, targeted sequencing analysis is performed using formalin-fixed paraffin embedded (FFPE) tissues. However, successful genetic analysis remains challenging because FFPE DNA is fragmented during the sample preparation process. Methods: Real-time PCR was performed to assess DNA quality and quantities. Targeted sequencing was performed using FFPE tissues fixed with different types of formalin. Results: DNA was less fragmented from samples fixed in low formalin concentration (10% vs. 20%) and neutral buffered conditions (neutral buffered vs. non-neutral). DNA fragmentation increased over the fixation time. In a preliminary test study, we compared fixation using 10% neutral buffered formalin (n = 180) and 20% formalin (n = 26). The success rate of targeted analysis was higher using 10% neutral formalin (98.3%; 177/180) compared with 20% formalin (34.6%; 9/26). In a validation study with additional formalin-fixed paraffin embedded tissues fixed with 10% neutral buffered formalin (n = 860), we reproduced these results and achieved a high success rate for targeted sequencing analysis (98.4%; 846/860). Conclusion: Our data show that 10% neutral buffered formalin is recommended for fixation of formalin-fixed paraffin embedded samples to achieve high success rate of targeted sequencing analysis.
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