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. 2012 Nov 13;2(11):e181. doi: 10.1038/tp.2012.101

Figure 1.

Figure 1

In vitro validation of lentiviral (LV)-siGAD67 virus. Image depicting primary cultures of hippocampal neurons from C57BL/6J mice (P10) infected with LV-SCR (a) or LV-siGAD67 (b). Overlay of fluorescence immunocytochemistry for GAD67 (red) in same field as in green fluorescent protein (GFP; green). Expression of GFP illustrates viral infectivity. These overlays illustrate the reduced expression of GAD67 signal from cells infected with LV-siGAD67 compared with LV-SCR. (c) Level of GAD67 expression was quantified by dividing the number of double label cells (GFP+GAD67) divided by infected cells (GFP). (d, e) Western blot processing of cells harvested from plate cultures revealed that LV-siGAD67-infected cells qualitatively expressed significantly less GAD67 protein than control-infected cells. (d) Western blot analysis of GAD67 protein levels from cell homogenates. Protein amounts determined from these western blots were normalized to levels of a GAPDH to loading control and expressed as percentages with reference to cells-alone control. (e) Digital image example of the western blot membrane showing protein bands of GAD67 and GAPDH protein in noninfected cells (cells alone), LV-SCR and LV-siGAD67-infected cells. Error bars denote 1 standard error of the mean (s.e.m.) Star indicates that the difference was statistically significant, P<0.05.