Figure 2. Validation of CDH23_V3 antisera. In A-C, arrows indicate untransfected cells; F-actin is stained in blue. A: A merged image showing HeLa cells 24 h after transfection with the expression vector GFP-Cdh23_v3a (see isoform designation Figure 1A). Cells were stained for CDH23_V3 with antibody TF648 (red) and F-actin antibody (blue). GFP fluorescence of transfected cells (green; B) overlapped with TF648 antibody staining (red; C) as revealed in the merged image (A). Scale bar equals 20 µm. D: A western blot was used to analyze protein extracts from HeLa cells transfected with GFP-Cdh23_v3a and untransfected controls. In the lysate of transfected cells a band of the expected size of roughly 50 kDa of the GFP-CDH23_V3a fusion protein (GFP approximately 27 kDa; CDH23_V3a approximately 26 kDa) was detected. E: Western blot of protein extracts from P60 mouse inner ear and retina revealed a band of roughly 26 kDa corresponding in size to CDH23_V3. β-actin was used as a loading control. Size standards are given in kDa.