A high-throughput Cognate Site Identity (CSI) microarray platform interrogating all 524 800 10-base pair variable sites is correlated to quantitative DNase I footprinting data of DNA binding pyrrole-imidazole polyamides. An eight-ring hairpin polyamide programmed to target the 5 bp sequence 5 '-TACGT-3 ' within the hypoxia response element (HRE) yielded a CSI microarray-derived sequence motif of 5 '-WWACGT-3 ' (W = A,T). A linear beta-linked polyamide programmed to target a (GAA)3 repeat yielded a CSI microarray-derived sequence motif of 5 '-AARAARWWG-3 ' (R = G,A). Quantitative DNase I footprinting of selected sequences from each microarray experiment enabled quantitative prediction of K-a values across the microarray intensity spectrum.