DiscussionThe transwell Boyden chamber has been the mainstay in vitro method for measuring directional migration/chemotaxis of cells for the last 30 years. However, it is widely acknowledged as technically tricky, hard to troubleshoot and frequently yields variable data. Here we introduce the new IncuCyte (TM) Chemotaxis System, designed to address these limitations and provide robust walkaway, fully kinetic and image-based assays in a 96-well format. The system is based around the novel ClearView 96-well plate consumable in which an ordered array of patterned 8uM microholes are created on a viewing surface in each well. Directional cell migration across the surface and toward chemoattractant placed on the underside of the plate is visualised over time using label-free IncuCyte (TM) live cell imaging and quantified with IncuCyte (TM) Chemotaxis Cell Migration image analysis software. This integrated solution is validated for both adherent and non-adherent cell types, including human neutrophils and T cells. The key benefits over transwells include full visualisation of the cell biology, low cell usage, easier workflows and relevant surface biology.