Evaluation of DNA/RNA quality from cell block of malignant mesothelioma and lung adenocarcinoma

Diagnostic Cytopathology 2022 April 5 [Link]

Shigenobu Tatsumi, Mao Takeuchi, Tomomi Fujii, Aya Sugimoto, Yoko Sekita-Hatakeyama, Takeshi Nishikawa, Tomoko Uchiyama, Kohei Morita, Hiroe Itami, Maiko Takeda, Masaharu Yamazaki, Chiho Ohbayashi


Malignant mesothelioma (MM) is a rare and highly lethal tumor that arises from mesothelial tissue on the surface of the chest and abdominal cavity. Cytological examination of body fluids, including pleural fluid and ascites, is essential for the differentiation of malignant mesothelioma from other carcinomas, such as lung and gastrointestinal carcinomas and metastatic tumors. To evaluate the effectiveness of cell block preparation procedures, which are used for immunocytochemical staining and genetic panel analysis of tumor-specific gene mutations, we used various fixatives. We also evaluated the effects of immunostaining, and the quality of nucleic acids for genetic analysis.

Methods: Cell blocks were prepared using the malignant mesothelioma cell lines MESO4 and H226 and non-small cell lung carcinoma cell line HCC78. The cells were fixed using 10% neutral buffered formalin and four different fixatives for liquid cytology. Fixed cells were formed into cell clusters using sodium alginate or centrifugation, and paraffin-embedded cell blocks were prepared.

Results: Cell blocks were morphologically evaluated by hematoxylin and eosin and immunocytological staining, and the nucleic acid quality was evaluated by DNA/RNA extraction, qPCR, and next-generation sequence analysis. D2-40 and WT1 staining differed depending on the fixation solution and the cell cluster formation method; however, the degree of nucleic acid degradation was not impaired by any method.

Conclusion: Although the morphological evaluation of cytology specimens is affected by the method of cell block preparation, it is still useful for nucleic acid extraction and gene panel analysis, as long as there are sufficient amounts of tumor cells.