(A-B), A tumor specimen is received and subjected to physical and enzymatic dissociation (A), yielding dissociated tumor tissue (B) containing spheroids, single cells, and macroscopic tumor. (C-D), This heterogeneous mixture is then sequentially applied to 100 μm and 40 μm filters (C) to obtain three separate fractions (D), S1 (>100 μm), S2 (40–100 μm), and S3 (<40 μm). E, the S2 fraction is pelleted and resuspended in collagen to be injected into the microfluidic culture device for subsequent ex vivo culture with indicated terminal readouts. Scale bars indicate 100 μm (D, E).