(1) choice of a solid support that is homogenous in size, suitable for nucleic acid synthesis, and compatible with the flow cytometer; (2) design, synthesis and purification of a range of modified nucleosides; (3) synthesis of the one-bead-one-compound library; (4) development of FABS methods for identification and isolation of high affinity modified aptamers; (5) tandem mass spectrometry to identify the position of type of the selected modifications; and (6) an EGFR binding assay (producing a K d ) to validate the affinity of the nucleoside modified aptamers for EGFR versus MinE07 (Fig. ). Step 1: Identification of solid support. A key requirement for FABS is use of uniform beads that can pass through the cytometer tubing and in front of the lasers as single beads. TentaGel® M NH₂ Monosized Amino TentaGel Microspheres (TG-beads) have a polystyrene backbone with a PEG spacer attached via alkyl linkage, which is resistant to acids and bases and so suitable for phosphoramidite chemistry. To be certain of microsphere size and uniformity, the diameters of a sample of TG-beads were measured using scanning electron microscopy (SEM), giving a mean bead diameter of 12.1 ± 0.7 µm (Figs. ). This was suitable for flow cytometry, as the tubing allows beads of up to 20 µm diameter to pass. Light and fluorescence microscopy also confirmed bead uniformity (Fig. ).