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. 2013 Jan 7;8(1):e53453. doi: 10.1371/journal.pone.0053453

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© 2013 Chellappa et al

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.

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293T cells were transfected with Flag-tagged SIAE or left untransfected as a negative control. The IgG heavy chain (IgG HC) was seen in all immunoprecipitated samples. (A) Cell lysates were immunoprecipitated with anti-Flag antibodies and analyzed by Western blotting using polyclonal anti-SIAE. An SIAE-GST fusion protein was run in the third lane as a positive control. (B) Cell lysates in both panels were immunoprecipitated with anti-SIAE, IgG and anti-Flag. The left panel displays the use of anti-SIAE for Western blotting. The immunoprecipitated samples in this panel reveal both overexpressed and endogenous SIAE. Immunoprecipitated proteins in the right panel were revealed by an anti-Flag Western blot. (C) Endogenous SIAE levels were further characterized in the B cell lines BJAB (left) and Ramos (right) by flow cytometric analysis. The data are all representative of three independent experiments.