FIG. 4.

Yop and V antigen expression and secretion in defined medium by lcrV yopJ Y. pestis KIM5-3241 complemented with pHT-V and pHT-VN68. Yersiniae pregrown at 26°C in the defined medium TMH containing (+) or lacking (−) 2.5 mM CaCl₂ were shifted to 37°C, and IPTG was added to cultures containing pHT-V and pHT-VN68 to induce expression of the complementing lcrV genes. After 5 h, cultures were harvested, and proteins in whole cells (C) and the cell-free culture medium (M) were analyzed in immunoblots probed with a mixture of antibodies specific for LcrV and YopE (A) or anti-YopM (B). Proteins were visualized by probing with alkaline phosphatase-conjugated secondary antibodies followed by development with NBT-BCIP.