Expression, purification, conversion to IgG1, and generation of stable clones Clones selected as described above were transformed into E.coli strain HB2151 for expression [36] . Briefly, a single clone was inoculated into 2YT supplemented with 100 units of ampicillin and 0.2% glucose and incubated at 37uC with shaking. When the OD600 reached 0.6-0.9, IPTG was added to achieve a final concentration of 1 mM and the culture was shifted to 30uC with shaking and incubated overnight. Cells were then collected, and lysed with polymyxin B (Sigma, St Louis) in PBS, and mixture subjected to Ni-NTA agarose bead (Qiagen, Hilden, Germany) purification. For IgG1 production, the heavy and light chains of the respective Fabs were cloned into the bi-cistronic expression vector pDR12 kindly provided by Dennis Burton (Scripps Research Institute, La Jolla, CA). For small scale IgG1 production, transient transfection and expression in Freestyle HEK 293F cells (Invitrogen, Carlsbad, CA) were used. For large scale production, stable clones were generated using CHO-K1 (ATCC, Manassas, VA) cells. Briefly, the heavy and light chains of the three human anti-plague IgG1s were cloned into pDR12 vectors and transfected into CHO-K1 cells. One day after transfection, the cells were replated and subjected to selection in GMEM medium supplemented with 25 mM MSX. Two weeks later, the MSX resistant clones were amplified further. The clones were tested for the expression of respective IgG1s and then adapted to growth in serum-free medium HyQSFM4CHO (HyClone, Logan, UT) supplemented with 30 mM MSX. The serum-free growth medium was then collected and passed through a protein A-sepharose resin column for IgG1 purification.