The walls were immunoblotted with the following main antibodies, mouse monoclonal antibodies directed against cleaved caspase 8 cytochrome C, p53 and bax, and rabbit polyclonal antibodies directed against ERK Gemcitabine structure, phospho ERK and JNK, and cleaved caspase 3, 9 and phospho JNK. The blot was then incubated with the corresponding anti mouse/rabbit immunoglobulin G horseradish peroxidase conjugated secondary antibody. Immunoreactive proteins were detected with the Enhanced Chemiluminescence Western blotting detection system. The relative density of the protein bands was scanned by densitometry using MyImage and quantified by Labworks 4. 0 software. HCT116, HT 29 colon cancer cells were plated in 24 well plates and transiently transfected with 0. 4 ug of the empty vector or the 100 nM of negative siRNA, DR4 or DR5 siRNA per well, using a mixture of plasmid and the WelFect EX PLUS reagent in OPTI MEM, according to manufacturers specification. Total RNA was extracted by RNeasy equipment. The RT reaction was performed using RNA to cDNA Kit. The PCR reaction was conducted with cDNA as a design Extispicy utilizing the primers below after an initial 1 min denaturation at 96 C, followed closely by the suggested cycles of 96 C for 1 min, 60 C or 63 C for 1 min and 72 C for 1 min. Generation of ROS was assessed by 2, 7 dichlorofluorescein diacetate, an oxidation painful and sensitive fluorescent probe. Intracellular H2O2 or low molecular weight peroxides may oxidize 2, 7 dichlorofluorescein diacetate to the highly fluorescent compound dichlorofluorescein. Briefly, cells were plated in 6 well plates, and subconfluent cells were subsequently handled with snake venom toxin for 30 min. After the cells were trypsinized, the 1×104 cells GW0742 were plated in 96 effectively plate and incubated with 10 uM DCFH DA at 37 C for 4 h. The fluorescence intensity of DCF was tested in a microplate reader at an excitation wavelength of 485 nm and an emission wavelength of 538 nm. The information were analyzed using the GraphPad Prism 4 ver. 4. April pc software. Data are presented as mean SD. The differences in all data were assessed by one of the ways analysis of variance. Once the G value in the ANOVA test indicated statistical importance, the differences were considered from the Dunnetts test. A value of r 0. 05 was considered to be statistically significant. To judge an effect of the snake venom toxin from Vipera lebetina turanica around the development of cancer of the colon cells, we analyzed the mobile viability by direct counting viable cells in Neubauer chamber. Snake venom toxin inhibited HCT116 and HT 29 cancer of the colon cell viability dose dependently. The IC50 values of snake venom toxin in HCT116 and HT 29 is 1. 14 ug/ml and 1. 24 ug/ml, respectively. Nevertheless, you will find no outstanding changes in CCD18 Co typical colon cell viability. To find out if the inhibition of cell viability by snake venom toxin was due to the induction of apoptosis, we evaluated the changes in the chromatin morphology of cells by using DAPI staining followed by TUNEL staining assays, and then a double labeled cells were analyzed by fluorescence microscope.