A statistical-based approach to assessing the fidelity of combinatorial libraries encoded with electrophoric molecular tags. Development and application of tag decode-assisted single bead LC/MS analysis

A statistical sampling protocol is described to assess the fidelity of libr aries encoded with molecular tags. The methodology, termed library QA, is b ased on the combined application of tag decode analysis and single bead LC/ MS. The physical existence of library compounds eluted from beads is establ ished by comparing the molecular weight predicted by tag decode with empiri cal measurement. The goal of sampling is to provide information on overall library fidelity and an indication of the performance of individual library synthons. The minimal sampling size n for library QA is 10x the largest sy nthon set. Data are reported as proportion (p) +/- lower and upper boundary (lb-ub) computed at the 95% confidence level (alpha = 0.05). As a practica l demonstration, library QA was performed on a 25 200-member library of sta tine amides (size = 40 x 63 x 10). Sampling was conducted three times at n similar to 630 beads per run for a total of 1902 beads. The overall proport ions found for the three runs were consistent with one another: p = 84.4%, lb-ub = 81.5-87.2%; p = 83.1%, lb-ub = 80.2-85.95; and p = 84.5%, lb-ub = 8 1.8-87.3%, suggesting the true value of p is close to 84% compound confirma tion. The performance pi of individual synthons was also computed. Corrobor ation of QA data with biological screening results obtained from assaying t he library against cathepsin D and plasmepsin II is discussed.