Rapid diagnosis of micrometastasis of gastric cancer using reverse transcription loop-mediated isothermal amplification

Abstract

Methods to detect metastases in biopsy specimens with certain rapidity and accuracy are essential to performing tailor-made surgeries for solid malignancies. Reverse transcription loop-mediated isothermal amplification (RT-LAMP) reaction is a novel technique for detecting mRNA expression of target sequences with high sensitivity and rapidity, even from crude samples without RNA purification. Applicability to detect lymph node (LN) micrometastasis of gastric cancer was tested. Total of 26 LNs were retrieved from 10 patients with primary gastric cancer. Each LN was serially sectioned, and every set of three serial sections were tested for routine histopathological (H&E) and immunohistochemical examination with anti-cytokeratin antibodies (IHC), and RT-LAMP analysis targeted cytokeratin 19 mRNA. Results from H&E/IHC and RT-LAMP analysis were compared in each set of sections. All the sections of those containing metastatic lesions equivalent to a volume of overt metastasis (maximum diameter >2 mm), 90% of those containing micrometastasis (between 2 and 0.2 mm) and 83% of those containing isolated tumor cells (<0.2 mm) were detectable using this procedure. Total analysis from lysates of clinical specimens required <75 min. This new technique is suggested to be an alternative to rapid diagnosis of micrometastasis based on conventional histopathological analysis.