Department of Pathology, Leiden University Medical Center, P.O.box 9600, L1-Q, Albinusdreef 2, 2300 RC Leiden, The Netherlands.
Exp Mol Pathol. 2013 Feb;94(1):121-5. doi: 10.1016/j.yexmp.2012.06.004. Epub 2012 Jun 29.
Preoperative biopsies or imbedded cytological cells will become more and more a primary source of tissue for molecular diagnostic analyses as a result of novel neo-adjuvant treatment regimens for several cancer types. Furthermore there is a growing need to examine metastatic cancer tissue. Hence, nucleic acids need to be reliably isolated and analyzed from small amounts of formalin-fixed and paraffin-embedded (FFPE) tissue. The limited numbers of (tumor) cells in these samples make high quality and sensitive DNA isolation challenging. Also demands for faster turnaround times are growing. Therefore, we evaluated a fully automated DNA/RNA isolation system and compared this with a manual, classical routine molecular pathology method. We compared the quality of the isolates from both tissue cores and micro-dissection for detection of hotspot mutations in KRAS, BRAF applying hydrolysis probe assays. In addition we determined whether the automated method decreases the hands-on-time and turnaround times in routine molecular pathology workflow. In conclusion, the automated method delivers high quality DNA from both small FFPE tissue cores and micro-dissected tissue material. In comparison to classical methods, less than 50% of starting tissue was sufficient as input for micro-dissection. Turnaround times decreased significantly and 50% less hands-on time was needed.
由于几种癌症类型的新型新辅助治疗方案，术前活检或嵌入的细胞学细胞将越来越成为分子诊断分析的主要组织来源。此外，越来越需要检查转移性癌症组织。因此，需要从少量福尔马林固定和石蜡包埋（FFPE）组织中可靠地分离和分析核酸。这些样本中的（肿瘤）细胞数量有限，使得高质量和敏感的 DNA 分离具有挑战性。对更快周转时间的需求也在增长。因此，我们评估了一种全自动 DNA/RNA 分离系统，并将其与手动的经典常规分子病理学方法进行了比较。我们比较了从两种组织芯和微切割物中分离的 KRAS、BRAF 热点突变检测的质量，应用水解探针检测法。此外，我们还确定了自动化方法是否减少了常规分子病理学工作流程中的人工操作时间和周转时间。总之，该自动化方法可从小的 FFPE 组织芯和微切割组织材料中获得高质量的 DNA。与经典方法相比，微切割所需的起始组织量减少了不到 50%。周转时间显著缩短，所需的人工操作时间减少了 50%。
