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. 2014 Sep 29;6(4):2012–2034. doi: 10.3390/cancers6042012

Figure 4.

Figure 4

Stat3β monoclonal antibodies are specific for the CT7 epitope found exclusively on Stat3β. Truncated pure Stat3α (amino acids 127–770), truncated Stat3β (amino acids 127–722) and truncated Stat3β devoid of CT7 (amino acids 127–717) were purified from bacterial supernatants as described in the Methods section. Decreasing amounts (100 ng, 30 ng, 10 ng, 3 ng, 1 ng, 0.3 ng) of truncated Stat3α (amino acids 127–770), truncated Stat3β (amino acids 127–722) were loaded onto consecutive wells and subjected to SDS-PAGE, transferred to nitrocellulose membranes and probed (A) with the three monoclonal antibodies 516 G10H9, 954 E9E7 and 1488 G6G5 as well as (B) Stat3α (Clone D1A5), two total Stat3 antibodies, one raised against a C-terminal peptide (clone 124H6) and one raised against a N-terminal peptide (aa 149–262, clone STAAD22A) and visualized by chemiluminiscence. Protein in a replica gel was also detected using silver staining and shown at the bottom of panel (B). In panel (C), decreasing amounts (100 ng, 30 ng, 10 ng, 3 ng, 1 ng, 0.3 ng) of truncated Stat3β (amino acids 127–722) and truncated Stat3β devoid of CT7 (amino acids 127–717) were loaded onto consecutive wells and subjected to SDS-PAGE, transferred to nitrocellulose membranes and probed with the three monoclonal antibodies 516 G10H9, 954 E9E7 and 1488 G6G5 as well as total Stat3 antibody (clone 124H6) and visualized by chemiluminiscence.