Figure 3

Purification of the P. pastoris -expressed ES,S ₀ , ES,S ₁ , ES,S ₂ , and ES,S ₄ , antigens. (A) Localization of the recombinant ES fusion antigens by Western blot analysis using mAb 24A12. Four ES fusion protein-expressing P. pastoris clones, co-expressing 0, 1, 2 and 4 copies of the S antigen (indicated by Arabic numerals on the right side of the immunoblot strips) were analysed. Aliquots of P. pastoris cultures were analysed before (U) and after methanol induction; induced cell extracts were analysed before (T) and after separation into supernatant (S) and pellet (P) fractions. Pre-stained protein markers were analysed in lanes ‘M’. Their sizes (in kDa) are indicated to the left. (B) Polypeptide profiles of the recombinant ES,S₀ (lane ‘0’), ES,S₁ (lane ‘1’), ES,S₂ (lane ‘2’), and ES,S₄ (lane ‘4’), antigens purified from the ‘P’ fractions of the induced cells (described in panel ‘A’), visualized by silver staining. (C) Western blot analysis of the purified proteins shown in panel ‘B’, using mAb 5S. Pre-stained protein markers were run in lane ‘M’. All other lanes are similar to panel ‘B’. For panels ‘B’ and ‘C’, the arrows on the left indicate the positions and sizes (in kDa) of the protein markers analysed in parallel; the arrows to the right indicate the positions of the ES (upper) and S (lower) antigens.