IHC in lung tissues of mice bearing B16F10 tumors was performed as described previously [19 (link)]. Briefly, formalin-fixed paraffin-embedded lung tissue was prepared as described for lung area measurement. We used anti-PCNA antibody (Chemicon-MAB424, 1:2500 dilution). Antibody binding was visualized using AEC (cat#3464, Dako, Glostrup, Denmark). Tissue samples were counter stained with hematoxylin according to standard methods.
IHC in lung tissues of mice bearing LLC1 tumors was performed by the Histopathology Core at the Koch Institute’s Robert A. Swanson Biotechnology Center using a Thermo Scientific LabVision 360 autostainer. Antigen retrieval was performed at 97 °C for 20 min in pH 6 citrate buffer (Abcam #3678). Slides were blocked using Rodent Block M (Biocare Medical #RBM961L). Primary antibodies were Cleaved Caspase 3 (CC3; Cell Signaling Technologies #9664 L, 1:800 dilution) and Ki-67 (Biocare Medical CRM 325B, 1:50 dilution). Secondary detection reagents were Mouse on Mouse HRP (Biocare Medical #MM620L) and Rabbit on Rodent HRP (Biocare Medical #RMR622L). After incubation with DAB Quanto Chromogen Substrate (ThermoScientific #TA-125-QHDX), slides were counterstained and scanned with Leica Aperio AT2.
IHC in lung tissues of mice bearing LLC1 tumors was performed by the Histopathology Core at the Koch Institute’s Robert A. Swanson Biotechnology Center using a Thermo Scientific LabVision 360 autostainer. Antigen retrieval was performed at 97 °C for 20 min in pH 6 citrate buffer (Abcam #3678). Slides were blocked using Rodent Block M (Biocare Medical #RBM961L). Primary antibodies were Cleaved Caspase 3 (CC3; Cell Signaling Technologies #9664 L, 1:800 dilution) and Ki-67 (Biocare Medical CRM 325B, 1:50 dilution). Secondary detection reagents were Mouse on Mouse HRP (Biocare Medical #MM620L) and Rabbit on Rodent HRP (Biocare Medical #RMR622L). After incubation with DAB Quanto Chromogen Substrate (ThermoScientific #TA-125-QHDX), slides were counterstained and scanned with Leica Aperio AT2.