Journal: PLoS ONE

Article Title: Avicin D, a Plant Triterpenoid, Induces Cell Apoptosis by Recruitment of Fas and Downstream Signaling Molecules into Lipid Rafts

doi: 10.1371/journal.pone.0008532

Figure Lengend Snippet: Proliferating Jurkat cells were exposed to different doses of avicin D for 24–72 h. A. Cell death was measured by trypan blue exclusion assay. The number of dead cells was counted as a percentage of total cells. B. Cell viability was measured by a CellTiter-Glo Luminescent Cell Viability Assay kit. Data are shown as mean±S.D. (n = 5). The number of surviving cells was counted as a percentage of total cells. C and D. Cell death and cell viability were measured in NB4 cells exposed to 0–4 µg/ml of avicin D for 24–48 h as described in the . Data are shown as mean±S.D. (n = 3). The number of dead or surviving cells was counted as a percentage of total cells. E. Proliferating Jurkat cells were exposed to 2 µg/ml of avicin D for 24 h. Cleavage of Caspase-8 induced by avicin D was analyzed by Western blot. 50 µg of total protein were loaded in each lane. α-Tubulin was detected to serve as an internal control. F. Time-dependent activation of Caspase-8 and apoptotic regulators in avicin D-treated Jurkat and NB4 cells. G. Time-dependent activation of Caspase-8 and apoptotic regulators in avicin D-treated U2OS cells.

Article Snippet: Cell death was examined by the cell death enzyme-linked immunosorbent assay (ELISA) according to the manufacturer's protocol of Roche Diagnostics (Pleasanton, CA) or by trypan blue exclusion kit from Sigma-Aldrich (St. Louis, MO) .

Techniques: Trypan Blue Exclusion Assay, Cell Viability Assay, Western Blot, Activation Assay

Journal: PLoS ONE

Article Title: Avicin D, a Plant Triterpenoid, Induces Cell Apoptosis by Recruitment of Fas and Downstream Signaling Molecules into Lipid Rafts

doi: 10.1371/journal.pone.0008532

Figure Lengend Snippet: (A & B) Proliferating Jurkat cells were pretreated with 2.5 mg/ml of MCD for 1 h, followed by 2 µg/ml of avicin D for 24 h. A. Cell death was quantified using a cell death ELISA showing enrichment of nucleosomes in the cytoplasmic fraction of Jurkat cells. Values represent the mean±S.D. (n = 3). B. Cell viability was determined by trypan blue exclusion assay. The number of surviving cells was counted as a percentage of total cells. C. MCD inhibits the activity of avicin D in a dose-dependent manner. Increasing concentrations of MCD were applied to Jurkat cells. One hour after treatment with MCD, 2 µg/ml of avicin D were added to the medium. Cell viability was determined by trypan blue exclusion assay after 24 h. Jurkat cells without treatment with MCD and avicin D were set as 100%. D. Jurkat cells were pretreated with 2.5 mg/ml MCD for 1h, followed by 2 µg/ml avicin D for 8 h. The cells were then fixed and stained with FITC-CTxB subunits to identify rafts (green fluorescence) and with anti-Fas antibody to identify Fas (red fluorescence). Area of colocalization between membrane rafts and Fas in the merge panels is yellow.

Article Snippet: Cell death was examined by the cell death enzyme-linked immunosorbent assay (ELISA) according to the manufacturer's protocol of Roche Diagnostics (Pleasanton, CA) or by trypan blue exclusion kit from Sigma-Aldrich (St. Louis, MO) .

Techniques: Enzyme-linked Immunosorbent Assay, Trypan Blue Exclusion Assay, Activity Assay, Staining, Fluorescence