A heterogeneous sample comprising a mixture of cells and cell media solution was loaded into a 1 ml syringe reservoir and each droplet was printed at pre-determined positions on a glass surface or into a 96-well plate.
Cells to be transfected were removed from the cultures and the CLONfectin/DNA media solution was gently applied.
CLONfectin/DNA- media solution was removed and then cells were washed with medium or 1× phosphate buffer saline (PBS).
The bottom agar layer contained 1.5 ml of 0.6% agar solution and 1.5 ml of media solution was poured and plated.
The GzmB/inhibitor media solution was transferred to wells that had been previously coated with FN and blocked as described above, before seeding of cells.
A 7.5% (wt/v) gelatin (Sigma) in media solution was made, which forms a solid at room temperature but melts at 37 °C.
The Casiopeína III-ia and the different aqueous media solutions were degassed before being loaded into the syringe and the reaction cell of the calorimeter.
All reagents, protein, and media solutions were prepared using water that had been passed through a Milli-Q ultrapurification system (Merck Millipore , Inc. to achieve a resistivity of 18 MΩ.
The media and selenomethionine solution were prepared according to the instruction supplied by the manufacturer.
All media and fixation solution were aspirated, and cells were washed once with 200 µL warm PBS++.
