The anti-GST-cHtrA fusion protein antibodies specifically detected the endogenous cHtrA produced by chlamydial organisms. The anti-cHtrA antibodies with or without absorption with GST fusion proteins were used to detect the endogenous proteins in C. trachomatis-infected cells (A) and on nitrocellulose membranes (B). (A) C. trachomatis-infected cells were processed for immunostaining as described in Figure 1A legend. Note that the antibody labeling of endogenous antigens was blocked only by corresponding but not unrelated control fusion proteins. (B) In a Western blot assay, HeLa alone or HeLa infected with C. trachomatis (Ct-HeLa), GST-CPAF & GST-cHtrA fusion proteins were used as antigens as indicated on top of the figure. The antigens blotted onto nitrocellulose membrane were detected with mouse antibodies as displayed at the bottom of the figure. The anti-CPAF mAb 100a is specific to the C-terminal fragment of CPAF (CPAFc) and the full length CPAF is rapidly processed into the N- and C-terminal fragments to form intramolecular dimmers for activity during chlamydial infection. The control antibodies anti-MOMP and anti-human HSP70 were used to indicate that the Ct-HeLa samples contain chlamydial organisms and both HeLa and Ct-HeLa samples were loaded with similar amounts. Note that each antibody only detected a major protein band migrated at the molecular weight that matched the corresponding chlamydial or host proteins as indicated on the right side of the figure.
Wu et al. BMC Microbiology 2011 11:87 doi:10.1186/1471-2180-11-87