Effective application of the methanol‐based PreservCyt™ fixative and the Cellient™ automated cell block processor to diagnostic cytopathology, immunocytochemistry, and molecular biology

Citations Over TimeTop 10% of 2013 papers

Abstract

We studied the feasibility of immunocytochemistry (ICC), in situ hybridization (ISH), and polymerase chain reaction (PCR) after Cellient(™) automated cell block processing, and tested whether methanol-based PreservCyt(™) fixation could replace formalin fixation, in an attempt to eliminate toxic formaldehyde vapors. Immunostaining with 30 different antibodies was performed on cell blocks from 73 FNA specimens and 42 body cavity fluid specimens prepared by Cellient(™) automated processing that uses the methanol-based fixative (PreservCyt(™) ). For each antibody we evaluated ICC in at least three different cell block specimens and compared it with immunohistochemistry (IHC) in formalin-fixed, paraffin-embedded (FFPE) histological sections from the corresponding tumors. The quality of DNA and RNA in Cellient(™) blocks was analyzed by ISH, applying a SYT gene break-apart assay and EBER probes, respectively. Moreover, DNA quality was analyzed by PCR by using primer sets for DNA products of 100, 200, 300, 400, 500, and 600 base pairs, and evaluated by gel electrophoresis. When compared with IHC results in corresponding FFPE tumor tissue from the same patient, 24 out of 30 antibodies showed concordant ICC results. With FISH, distinctive hybridization signals were observed for SYT DNA sequences and EB virus RNA sequences. With PCR, DNA products, up to 600 base pairs in size, were readily observed after gel electrophoresis. The antibodies that showed concordant immunostaining in Cellient(™) blocks could be applied to diagnostic algorithms that proved to be helpful in the discrimination of major tumor types (carcinoma, lymphoma, melanoma, and germ cell tumors), discrimination of carcinoma subtypes, and determination of primary tumor site in cases of metastatic carcinoma. In a separate study, we found that the application of ICC to this cell block technique provided additional diagnostic and clinically important information in 24% of 100 consecutive cases. The high quality of DNA and RNA in Cellient(™) cell blocks allowed sensitive and specific molecular biologic analysis, in particular FISH and PCR.

Related Papers

• → Using Kryofix as Alternative for Formalin Results in More Optimal and Standardized Immunostaining of Paraffin Sections(1992)29 cited
• → Comparison of cellular composition of induced sputum analyzed with Wright staining and immunocytochemistry(2001)15 cited
• → A comparison study of histochemical staining of various tissues after Carnoy’s and formalin fixation(2011)5 cited
• Formalin is deleterious to cytoskeleton proteins: do we need to replace it by formalin-free Kryofix?(1991)
• 여려 염색조건이 AgNORs 염색에 미치는 영향(1993)