The original MinE07 aptamer was synthesised, along with the three hit modified aptamers, which each had an additional biotin at the experimentally identified truncated length (MinE07UA-Biotin, MinE07UB-Biotin and MinE07UC-Biotin) and also the expected full-length versions of the hits (MinE07UA'-Biotin, MinE07UC'-Biotin) for validation (S4, Figs. 127-141). Control aptamers MinE07-U-Ph-Biotin, MinE07-U-Vi-Biotin and MinE07-U-I-Biotin, which have uniform modification of all their uridines (comparable with the state of the art for modified aptamers) were also synthesised (S4, Figs. ). The strands were purified by polyacrylamide gel electrophoresis, and confirmation of the full-length product was obtained by performing a 4'-hydroxyazobenzene-2carboxylic acid (HABA) assay (S4, Fig. ). This reports the presence of biotin, which can only exist in full length strands as it is the final (5') monomer, and the capping step in the synthesis would prevent its addition to truncated strands. HABA can be displaced from the binding site of streptavidin by biotin. Protein A plates (96-well) were incubated with EGFR-Fc, after which aptamers were added followed by streptavidin-alkaline phosphatase (AP) and an AP chemiluminescent substrate. In the absence of biotin, HABA quenches chemiluminescence via FRET. All the newly synthesised aptamers gave a positive signal and were carried forward for K d determination in an ELISA-based EGFR binding assay.