Image studio software 4

Westernblot images in Figs. 1, 4 and 6 were acquired with the Image Studio Software 4.0 (LICOR) with the scanning intensity of 5.0 for channel 700 and of 4.5 for channel 800.
Immunofluorescence images in Fig. 2 were taken with the Leica DMI4000B microscope with the camera Leica-DFC3000G-0001062113 and the objective N PLAN L 20×/0.35 DRY using the Leica Application Suite X software in the red, green and blue channel. The acquisition was performed with the time and space resolution XY = 0.959 µm and Z = 4.490 µm, 12 bit images were taken with a resolution of 1296 × 966 pixels. All pictures were taken with the same settings. In the illustrated TUNEL and Ki67 pictures brightness was increased by 30% for all conditions in the Power Point software. Images in Fig. 3 were taken with the same microscope and the same settings only for the green channel, brightness was increased by 20% for all conditions in Power Point software.
Images in Fig. 5 were taken with the same microscope and camera and the objective HC PL APO 40×/0.85 DRY. The acquisition was performed with the time and space resolution XY = 0.395 µm and Z = 0.761 µm, 12 bit images were taken with a resolution of 1296 × 966 pixel. All pictures were taken with the same settings. Colour saturation was increased to 200% for all conditions in Power Point software.

To compare the expression of the livin isoforms α and β in ACT and in adjacent NAG and in ACC cell lines, we additionally performed regular RT-PCR followed by size differentiation agarose electrophoresis. The primers used amplify a fragment including the 5′-part of exon 6, deleted in livin β [22 (link)] (Supplementary Table 2B). The expression of endogenous β2-microglobulin [18 (link)] was used for normalization. Primers were purchased from Eurofins Genomics (Ebersberg, Germany). RT-PCR was performed using the Sybr Green PCR Master Mix (Life Technologies, Warrington, UK). PCR products were identified by 4% agarose gel electrophoresis. We differentiated between two fragment sizes: 216 bp band for livin α and 162 bp for livin β. After gel documentation (Biometra, Göttingen, Germany) a semi-quantitative evaluation of the bands was performed using the Image Studio software 4.0 (LI-COR, Bad Homburg, Germany). Hela cells were used as positive control [13 (link)].