Figure 2.
Purification of bTAP from murine milk. (A) RP-HPLC of whey fraction of murine milk purified on a C18 SepPak column and eluted with 60% acetonitrile. The eluate was applied to a RP-HPLC column and fractionated with a linear gradient of 0–60% acetonitrile in H2O/0.1% trifluoroacetic acid. The upper plot is nontransgenic milk alone, while the lower plot is similar material with bTAP isolated from tracheal mucosa added. The arrow indicates an additional peak from bTAP addition. (B) Transgenic (TG) versus nontransgenic (NTG) milk purified as in A. The peak of increased size at 32.5 min corresponds to antimicrobial activity in an in vitro bacterial lysis assay (data not shown). (C) IEC of material corresponding to 32.5-min fraction of HPLC shown in B. Antimicrobial activity in the in vitro assay is present in the 30-min fraction of the TG but not the NTG sample (data not shown).