Studio lite software

To investigate the expression of Gα_i proteins in human pituitary tumors, Gα_i-1, Gα_i-2 and Gα_i-3 immunostainings were performed in four prolocatinomas, six somatotropinomas, three ACTH and four NFPA tumors. All tumors were AIP mutation negative. Antibodies used were mouse monoclonal antibody against Gα_i-1 (SPM397, sc-56536, Santa Cruz, 1: 40), rabbit polyclonal antibody against Gα_i-2 (T19, sc-7276, Santa Cruz, 1: 60) and mouse polyclonal antibody against Gα_i-3 (H00002773-B01P, Abnova Corp. Taipei city, Taiwan, 1: 50). Anti-mouse/rabbit/rat secondary antibody, Poly-HRP-GAM/R/R (DPVB55HRP, Immunologic, Duiven, Netherlands) and DAB chromogen (Lab Vision Corporation, Fremont, CA, USA, Thermo Fisher Scientific, Watham, MA, USA) were used for detection. Immunostaining protocol was applied as described [30] (link). The staining intensities of Gαi proteins were scaled as negative (0), weak (1), moderate (2), or strong (3). The images were taken and edited by Leica DM LB microscope (Meyer Instruments, Houston, TX, USA), Olympus DP50 camera (Olympus Corporation, Tokyo, Japan) and Studio Lite software (Licor, Lincoln, NE, USA).

Histology was performed on 4 mm sections collected at different time points, formalin-fixed, paraffin-embedded tissues for hematoxylin-eosin staining, and β-galactosidase staining. Images were acquired using an Olympus BX51 microscope (Olympus) and Studio Lite software (Li-COR Biosciences, GmbH).
The hematoxylin-eosin staining was performed by means of a Leica ST5020 (Barcelona, Spain) and CV5030 automatic dye and mounter. To this end, the preparations posed as successive solutions of xylene, alcohols in decreasing gradation (100, 96, and 70%) to hydration, Carazzi hematoxylin, running water, eosin and 1%, increasing alcohol solutions up to complete dehydration (70, 96, and 100%) to finish with the rinsing in two xylene baths and assembly with permanent medium (Sigma-Aldrich).
Immunohistochemistry was performed on 4-μm sections of formalin-fixed, paraffin-embedded tissues. The antibody used was anti-human NIS (hNIS) (rabbit polyclonal) (1:2000), kindly donated by Dr. de la Vieja.

To quantitatively compare the intensity of Coomassie-stained protein bands (Fig. 4A) or signals obtained by detecting specific proteins with western blotting (Fig. 6B), the LiCOR Studio-Lite software was used.