Figure 4

Innate tumour cell selectivity is exhibited by the recombinant histone H3 protein and is further enhanced by the Apoptin-tNTS. A. SR5 (normal) or SAOS-2 (tumour) cells were incubated for 4 hours in media containing 1.17 μM indicated recombinant protein. Cells were then washed with warm 37°C PBS and treated with DMEM lacking phenol red, containing 1.25 μg/mL Hoescht stain for 30 min. Cells were thoroughly washed in warm PBS and imaged using CLSM as per mammalian cell transfection (above). The blue channel shows Hoescht stained nuclei and the green channel shows the uptaken GFP-tagged recombinant protein. Blue (Hoescht) and Green (GFP) channels are presented separately in grayscale and merged in colour for clarity. B. Analysis of digitised images such as those in A was used to determine the relative mean fluorescence in the nucleus of cells as described in Materials and Methods. The mean fluorescence value was then normalised to the GFP value representing background fluorescence in each cell line. C. The data in B was used to calculate CCI values for GFP-H3 and GFP-H3tNTS recombinant proteins, relative to GFP alone (as disregarded background). Results are from a single typical experiment from a series of two similar experiments.