We saw an abolishment from the selection of the poor hit cP4_2 and the peptide hit without the warhead cP4_3. Two further peptides that were uniquely identified in this second screen, cP4_13 and cP4_18, were synthesised. Both were shown to covalently bind to C645 of PADI4 by intact MS (Figure ). cP4_13 closely resembled cP4_2 from the first selection, but with only one warhead 2 present. Neither peptide had strong inhibitory activity against PADI4 (Figure ) and cP4_13 was considerably less active than cP4_2. Characterisation by SPR also showed that these peptides were among some of the poorest binders synthesised (Figure ). These results showed that the alternative selection conditions did not eradicate the least potent hits, however their enrichment levels were lower relative to the more active cP4_4, cP4_7 and cP4_10, which might have helped with initial peptide selection for synthesis by SPPS.