RESUMEN
AIM OF THE STUDY: One of the critical steps in molecular oncology diagnostics is obtaining high quality genomic DNA. Therefore, it is important to evaluate and compare the techniques used to extract DNA from tissue samples. Since formalin-fixed, paraffin-embedded (FFPE) tissues are routinely used for both retrospective and prospective studies, we compared three commercially available methods of nucleic acid extraction in terms of quantity and quality of isolated DNA. MATERIAL AND METHODS: Slides prepared from 42 FFPE blocks were macro-dissected. Resulting material was divided and processed simultaneously using three extraction kits: QIAamp DNA FFPE Tissue Kit (QIAGEN), Cobas DNA Sample Preparation Kit (Roche Molecular Systems) and Maxwell 16 FFPE Plus LEV DNA Purification Kit (Promega). Subsequently, quantity and quality of obtained DNA samples were analysed spectrophotometrically (NanoDrop 2000, Thermo Scientific). Results of quantitative analysis were confirmed by a fluorometric procedure (Qubit 3.0 Fluorometer, Life Technologies). RESULTS: The results demonstrated that the yields of total DNA extracted using either Maxwell or Cobas methods were significantly higher compared to the QIAamp method (p < 0.001). The Maxwell Extraction Kit delivered DNA samples of the highest quality (p < 0.01). However, the highest total yield of extracted DNA was achieved with the Cobas technique, which may be due to a higher volume of eluate compared to the Maxwell method. CONCLUSIONS: To our knowledge, this is the first paper which directly compares three extraction methods: Cobas, Maxwell and QIAamp. The data herein provide information required for the selection of a protocol that best suits the needs of the overall study design in terms of the quantity and quality of the extracted DNA.
RESUMEN
AIM OF THE STUDY: To determine the correlation of protein serum levels of two cytokines and their polymorphisms, which have an influence on their expression. MATERIAL AND METHODS: The study group consisted of 65 patients (33 men, 31 women) who met the criteria for inclusion and exclusion of pancreatic cancer, and 41 patients (25 men, 16 women) with colorectal cancer. The control group consisted of 100 healthy volunteers (63 men, 37 women). Detection of polymorphisms was performed using TaqMan probes, and concentration of proteins by ELISA method. RESULTS: The mean TNF-α concentration in patients with colorectal cancer was significantly higher compared to the control group, p< 0.0001. A statistically significant difference was noted when comparing both study groups, p = 0.0009. The analyses show that the occurrence of the polymorphic genotype -308AA of the TNF-α gene was not correlated with the increased concentration of the examined protein in patients with both pancreatic and colorectal cancer. It was also noted that the concentration of TGF-ß protein was significantly higher in patients with colorectal cancer than in patients with pancreatic cancer. These results also proved to be statistically significant, p = 0.0353. CONCLUSIONS: The only statistically significant effects were the correlations between patients belonging to a specific group (pancreatic cancer/colorectal cancer/control) and average protein levels. There was no effect of sex or genotype on the occurrence of elevated levels of TNF-α and TGF-ß protein control, despite their variability in particular types of cancer.
RESUMEN
The pancreatic carcinoma is a leading cause of death in cancer carriers worldwide. The early diagnostic is difficult due to late stage during diagnosis, lack of characteristic symptoms and also multifactor basis. In cancer development take part both, environmental and genetic factors, alone or in conjunction with each other. The nonspecific biomarkers of cancers are a reason for the search for more accurate factors which allow for fast and personalized diagnostics. Some of cancers have identified molecular (metabolic, biochemical or genetic) markers but in most cases the only clue is patient`s interview and abnormal levels of organ functions markers. Possible genetic basis of cancer suggests to widen studies on connection between environmental factors with both, nuclear and mitochondrial, genes changes.