Jikuzono Tomoo, Horikawa Aya, Ishikawa Tomoko, Hirokawa Mitsuyoshi, Sugitani Iwao, Inui Takashi, Ishibashi Osamu
Laboratory of Biological Macromolecules, Department of Applied Life Sciences, Graduate School of Life & Environmental Sciences, Osaka Prefecture University, 1-1 Gakuen-cho, Sakai, 599-8531, Japan.
Department of Endocrine Surgery, Kanaji Thyroid Hospital, 1-5-6 Nakazato, Kita-ku, Tokyo, 114-0015, Japan.
BMC Res Notes. 2018 Nov 20;11(1):822. doi: 10.1186/s13104-018-3914-4.
Fine-needle aspiration biopsy (FNAB), an important diagnostic tool given its simplicity, safety, and cost-effectiveness, is fast becoming a popular procedure in the diagnosis of thyroid diseases. Generally, cells isolated from biopsies are transferred directly to microscope slides to prepare smears for cytopathological examination; however, the technical difficulties of this procedure often cause poor reproducibility, which limits the accuracy of diagnostic results. Liquid-based cytology (LBC), in which isolated cells are collected in a fixative solution, is advantageous in that it facilitates the preparation of homogenous cytological specimens. However, LBC has not been applied to molecular diagnoses, such as RNA expression-based diagnosis, mainly because of difficulties in cell recovery and RNA isolation. This study was aimed to improve RNA extraction from papillary cancer-derived K1 cells and thyroid FNAB specimens suspended in LBC solutions.
K1 cells suspended in CytoRich-Red and CytoRich-Blue, fixatives for LBC, were efficiently recovered by trapping to glass-fiber filters. Importantly, subsequent Proteinase K treatment was essential for efficient RNA extraction from the fixed cells. This finding was also applicable to RNA extraction from CytoRich-Red-fixed thyroid FNAB specimens processed in the same way. Consistently, U6 small nuclear RNA was detected in these RNA samples by reverse transcription-polymerase chain reaction.
细针穿刺活检（FNAB）因其操作简单、安全且具有成本效益，是一种重要的诊断工具，在甲状腺疾病诊断中迅速成为一种常用方法。一般来说，从活检中分离出的细胞直接转移到显微镜载玻片上制备涂片用于细胞病理学检查；然而，该操作的技术难点常导致重复性差，从而限制了诊断结果的准确性。液基细胞学（LBC）是将分离出的细胞收集在固定液中，其优点在于便于制备均匀的细胞学标本。然而，LBC尚未应用于分子诊断，如基于RNA表达的诊断，主要是因为细胞回收和RNA分离存在困难。本研究旨在改进从悬浮于LBC溶液中的乳头状癌来源的K1细胞和甲状腺FNAB标本中提取RNA的方法。
悬浮于LBC固定液CytoRich-Red和CytoRich-Blue中的K1细胞通过捕获到玻璃纤维滤膜上能有效回收。重要的是，随后的蛋白酶K处理对于从固定细胞中高效提取RNA至关重要。这一发现同样适用于以相同方式处理的CytoRich-Red固定的甲状腺FNAB标本的RNA提取。通过逆转录-聚合酶链反应在这些RNA样本中一致检测到U6小核RNA。
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