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. 2017 Jan 5;24(2):92–103. doi: 10.1038/gt.2016.80

Figure 1.

Figure 1

TMZ-CD40L is membrane-bound post expression. (a) Schematic figure of TMZ-CD40L compared with wild-type CD40L. (b) 293 cells were transfected with a TMZ-CD40L-containing plasmid (LP-700; red line), empty plasmid (LP(−); blue line) or untransfected (green broken line) and analyzed day 2 by flow cytometry. (c) 293 cells transfected with LP-700 or LP(−) were lysed at day 2 post transfection and the protein suspension analyzed by gel electrophoresis and western blot to detect TMZ-CD40L. Samples boiled in reducing sample buffer (R) or not boiled but mixed with a non-reducing sample buffer before gel electrophoresis. (d) Pancreatic cancer cell lines were untransduced or transduced with LOAd viruses transferring TMZ-CD40L (LOAd700), empty virus LOAd(−) or with AdCD40L-expressing wild-type CD40L. At day 2, the cells were analyzed for CD40L expression using flow cytometry. Black line demonstrates CD40L expression in LOAd700- or AdCD40L-transduced cells compared with LOAd(−) cells (filled curve). A representative figure of transduced PaCa3 cells is shown. (e) The supernatants were collected from the cell cultures in d and the presence of soluble CD40L (sCD40L) was determined by ELISA. The difference of sCD40L levels between the groups was determined by the Student's t-test with Welsh correction (P=0.02) and the error graphs show s.e.m.