The cell culturemediumwas DMEM for growing SK N BE2 cell line and was RPMI 1640 for growing SH SY5Y cell line. Previous todrug treatment, cellswere growntill 80%confluency and then starved within their respective cell culturemediumcontaining 14 days FBS for 24 h. The Bcl 2 chemical MK-2206 molecular weight and genistein were obtained. Drugs were dissolved in dimethyl sulfoxide tomake a stock option and aliquots were stored at 20 C until ready for use. Doseresponse studies were conducted to ascertain the amounts of the medications for induction of apoptotic death. Cell viability was determined using an MTT colorimetric assay system. The fundamental principle with this assay is to gauge the activity of mitochondrial enzyme system that changes yellow MTT to purple colored formazan. SH SY5Y cells and both SK Deborah BE2 were seeded at 3?105 cells/well in two 96 well plates separately. Different doses of GST and HA and their combination were added to each plate in triplicates and plates were incubated over night in a humidified incubator containing 550-watt CO2 at 37 C. Then, MTT reagent was added in each plate and incubated for 4 h at 37 C. Formazan precipitate was dissolved by pipetting each well up and down with 100 ul of isopropyl alcohol. Plates were read on the spectrophotometer using 570 nm since the test wavelength. Cell viability data were analyzed using CompuSyn pc software to determine a combination index for synergism in drug combination studies. Traditionally, CI 1 indicates antagonism, CI_1 Gene expression indicates chemical effect, and CI 1 indicates synergism in the effective doses. We observed a low CI using 10 uM HA 250 uM GST in SK Deborah BE2 cell line and also a low CI using 5 uM HA 100 uM GST in SH SY5Y cell line. Therefore, these specific doses of the drugs and their combinations were chosen because of their complete inhibitory action on cell growth in all other experiments. Both cell lines in culture dishes were treated with HA, GST, and HA GST for 24 h and examined under the phase contrast microscope. purchase CX-4945 Treatments caused various morphological features of apoptosis in cells on the plates. Using phase contrast microscopy, black and white pictures were taken. More, cells from each treatment were washed with PBS and sedimented on slides using the Eppendorf 5804R centrifuge at 106 g for 5 min. Cells were stained with Wright discoloration and fixed with 95% ethanol. Morphological characteristics of apoptotic cells were detected underneath the light microscope. Morphological features of apoptosis included reduction of existence of membranebound apoptotic bodies, and cell volume, chromatin condensation. Four randomly selected areas were measured for at least 800 cells. The percentage of apoptotic cells was determined from three split up studies.