Induction of apoptosis In order to examine the aftereffect of the anti apoptotic Bcl 2 expression, the cells were subjected to 800 ugml hygromycin B in DME medium supplemented with 10% FBS or were deprived of FBS in DME medium and then a possibility was decided. Detection of DNA ladder Apoptosis was dependant on measuring the degree of DNA fragmentation in an agarose gel. Cells were harvested from tradition, lysed by vortexing vigorously in Tris HCI pH 7. 4 and EDTA 1 mM buffer containing 0. A day later Triton X 100, and centrifuged for 10 min at 16,OOOxg Capecitabine solubility at 4 C. The supernatant was treated with RNase A for 60 min at 37 C and then ethanol and sodium acetate were traditionally included. After centrifugation, the DNA pellets acquired were redissolved in TE buffer and packed on 1% agarose gels for electrophoresis. DNA molecular weight markers and dna bands were visualized by staining with ethidium bromide. Determination of albumin production The albumin concentration in the tradition supematant was determined by ELISA. The ELISA was performed in 96 well plates. The wells were first covered with goat anti human albumin antiserum and plugged with skim milk. Consequently, the conventional wells were coated with purified human serum albumin. The others of the wells were lined with the fresh samples. Finally, the wells were covered with a horseradish peroxidaseconjugated anti human Lymphatic system albumin polyclonal antibody and an answer of o phenylenediamine in citric acid buffer was added to the wells. The absorbance was read at 490 nm. Measurement of ammonia removal On each day the culture supernatant was removed and sold for fresh medium supplemented with I mM ammonium chloride. The decrease in the me dium focus through the initial 2 h was determined by measuring the levels in medium examples. The ammonium in the medium samples was measured utilizing an ammonia test in line with the indophenol method. Measurement of cytochrome P450 activity The inducibility PFI-1 concentration of cytochrome P450 CYPlA enzyme activity was assessed by measuring ethoxyresorufin O deethylase activity after treatment in tradition with 2 PM 3 methylcholanthrene for 24 h. After day 2, the culture medium was removed daily and traded for serum free DME medium containing 1OmM ethoxyresorufin as a, and 10 pM dicumarol to be able to prevent further metabolism of the resorufin produced. After incubation for just two h at 37 C, the culture supernatant was collected and centrifuged. Supernatant fluorescence was measured having an 850 Fluorescence Spectrophotometer at 585 nm emission and 530 nm excitation. The BCMG bcl 2 neo vector was selected in the current presence of G418 and introduced into human hepatoblastoma HepG2. By decreasing dilution, one clone was called and obtained HepG2 Bcl2.