Generation of a specific fluorescence in situ hybridization test for the detection of ovarian carcinoma cells

Objective Investigations of ovarian carcinoma (OvCa) cells require identification of tumor cells. Array-based, comparative genome hybridizations on 30 OvCa identified three genomic loci (8q24.23; 17p12; 18q22.3), each is not diploid in OvCa up to 60%. A fluorescence in situ hybridization (FISH) probe of these three loci is intended to identify tumor cells by their signal pattern deviating from a diploid pattern.

Material and methods Human DNA from these three loci is isolated from bacterial artificial chromosomes, amplified and labeled with fluorescent dyes. Dropped cells are pretreated with a 0.005% pepsin and 1% buffeted formaldehyde. After a standard FISH procedure, 88 OvCa suspensions from primary tumors, three OvCa cell lines, three lymphocyte suspensions and one mesenchymal human cell line LP-3 are analyzed by fluorescence microscopy.

Results On average, 15% of the lymphocytes deviate from the expected diploid signal pattern. The standard deviation (SD 7%) is added three times to the mean, giving a limit value of 36%. If this value is exceeded, tumor cells are detected. The mesenchymal cell line LP-3 shows only 21% as a negative control. All OvCa cell lines exceed this value as positive controls. In 88 OvCa primary cultures, six cases fall below this cut-off as false negatives. This results in a sensitivity of 0.935 for this FISH test for the detection of tumor cells in cell suspensions.

Summary This FISH test allows the identification of cells with genomic imbalances, as can be expected in OvCa, and thus an identification of the tumor cell in experimental approaches.

Publikationsverlauf

Artikel online veröffentlicht:
11. Oktober 2022