Figure 2
Protein and mRNA expression analysis of AK2 in cell lines derived control (CT) and patient (PT). (A) Fibroblast cells from CT and PT were analyzed for the presence of AK2 antigen by SDS-PAGE followed by western blotting with anti-AK2 and anti-β-actin antibodies. Twenty-five µg protein from fibroblasts’ lysates were loaded for both CT and PT (see Supplementary Figs S1 and S2 for full blot). (B,C) Immunofluorescence staining of fibroblasts and bone marrow from CT and PT. AK2 antigen was visualized with green fluorescently tagged antibodies and mitochondrial cytochrome c oxidase subunit 1 (MTCO1) was visualized with red fluorescently tagged antibodies. Nuclei were visualized with DAPI staining. The merged image shows co-localization of AK2 and MTCO1 in mitochondria as yellow/orange in the CT fibroblasts while there is essentially no green staining that would show co-localization with MTCO1 in the patient fibroblasts. Scale bar, 50 μm. (D) RT-PCR for AK2 and GAPDH on CT, PT and negative control (H2O) samples. GAPDH was used as a standard reference gene. Both CT and PT PCR reactions resulted in similar cDNA band instensity (see Supplementary Fig. S3 for full image of agarose gel electrophoresis).
