Preparation of hPSC secretions: Secretions from hPSCs (n=5) that had been standardised to 2000 pg HGF ml−1 were pretreated for 1 h at 37 °C with 60 μg ml−1 AMG102 (HGF-neutralising antibody; Amgen, Thousand Oaks, CA, USA), 60 μg ml−1 IgG (isotype control for AMG102; Amgen), 300 μg ml−1 gemcitabine (Hospira, Mulgrave, VIC, Australia) or combination of gemcitabine and AMG102. These concentrations of treatment compounds were calculated corresponding to in vivo dose used in these studies (see Supplementary Methods ).
Assessment of AsPC-1 cell proliferation, migration and apoptosis: The effects of HGF inhibition on AsPC-1 proliferation were measured using the Cell Counting Kit-8 (Dojindo Molecular Technologies, Rockville, MD, USA). AsPC-1 cell migration was assessed using a modified Boyden chamber method as published by us previously using cell culture inserts with a porous membrane at the bottom (8-μm sized pores) (Becton-Dickinson, Bedford, MA, USA) placed in 24-well culture plates. Apoptosis was assessed by Annexin V staining. The above methods are described in detail in theSupplementary Methods .
Assessment of AsPC-1 cell proliferation, migration and apoptosis: The effects of HGF inhibition on AsPC-1 proliferation were measured using the Cell Counting Kit-8 (Dojindo Molecular Technologies, Rockville, MD, USA). AsPC-1 cell migration was assessed using a modified Boyden chamber method as published by us previously using cell culture inserts with a porous membrane at the bottom (8-μm sized pores) (Becton-Dickinson, Bedford, MA, USA) placed in 24-well culture plates. Apoptosis was assessed by Annexin V staining. The above methods are described in detail in the