Cell-based extracellular protein-protein interactions. (A) COS7 cells transfected with either HNT or OPCML expression constructs were fixed and then incubated with NEGR1-hFc bait protein. The cells were then washed and stained with anti–human AF488 (green) and scanned on the IN Cell Analyzer 2000. The same reagents for transfection and staining were used on a standard 384-well microtiter plate (Aurora, Brooks Automation) and a DropArray plate for a side-by-side comparison of the 2 formats. Hoechst staining (blue) of the nuclei shows the same cell density on both plate formats. Scale bar = 70 μm. (B) Quantification of the mean intensity of the green channel from the images acquired. With the use of 3 different cell types, equivalent intensity values are seen when NEGR1-hFc binds to cells expressing 3 different expression constructs of known NEGR1 binding partners. The same values are generated from the standard 384-well format, but only for the adherent cell line (COS7), whereas the weakly adherent (293T) and suspension adapted (293S) cell lines were not retained during the washing procedure on the standard 384-well plate format.