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. 2003 May;10(3):459–468. doi: 10.1128/CDLI.10.3.459-468.2003

FIG. 2.

FIG. 2.

Expression of recombinant P46 and P65c and Western blotting patterns of specific monoclonal antibodies. (A) Expression of the GST-P46 and GST-P65c recombinant fusion proteins in E. coli. Lane 1, IPTG-induced GST recovered from pGEX-4T1-transformed bacteria (GST = 26 kDa); lane 2, purified GST-P46 recombinant fusion protein (72 kDa); lane 3, cleaved recombinant GST-P46 fusion protein (46 kDa); lane 4, purified GST-P65c recombinant fusion protein (70 kDa); lane 5, cleaved GST-P65c recombinant fusion protein (44 kDa); lane 6, noninduced recombinant bacteria; lane M, Rainbow molecular mass markers. (B) Polypeptide specificities of pooled MAbs produced against E. coli-expressed P46 and P65c recombinant proteins. The reactivities of pooled MAbs were tested against the fusion and cleaved recombinant proteins. Lane 1, IPTG-induced pGEX-4T1-transformed bacteria (GST = 26 kDa); lane 2, purified GST-P46 recombinant fusion protein (72 kDa); lane 3, cleaved recombinant GST-P46 fusion protein (46 kDa); lane 4, purified GST-P65c recombinant fusion protein (70 kDa); lane 5, cleaved GST-P65c recombinant fusion protein (44 kDa); lane 6, noninduced recombinant bacteria; lane M, Rainbow molecular weight markers. (C) Reactivities of the anti-P46 and anti-P65c MAbs as determined by Western blotting using whole M. hyopneumoniae lysate as antigenic preparation. The figure illustrates the reactivity patterns as follows: lane 1, polyclonal antiserum to M. hyopneumoniae; lane 2, MAb 7D3-E9 (anti-P46); lane 3, MAb 7D3-C11 (anti-P46); lane 4, MAb 6A4-G9 (anti-P46); lane 5, MAb 4D11-C11 (anti-P65c); lane 6, MAb 1D3-C6 (anti-P65c); lane 7, MAb 4D11-G8 (anti-P65c).