Detection and quantification of Panton-Valentine leukocidin in Staphylococcus aureus cultures by ELISA and Western blotting: diethylpyrocarbonate inhibits binding of protein A to IgG.

Hien M Nguyen, Miguel A Rocha, Koteswara R Chintalacharuvu, David O Beenhouwer

Division of Infectious Diseases, Veterans Affairs Greater Los Angeles Healthcare System, Los Angeles, California, United States.

Journal of immunological methods 2010 Apr 30

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Enzyme-linked immunosorbent assay (ELISA) and Western blotting are common techniques used to detect and quantify proteins in Staphylococcus aureus culture supernatants, such as Panton-Valentine leukocidin (PVL). However, protein A (Spa) secreted by most S. aureus strains may interfere with these assays by binding to the capturing and detecting antibodies. Here, we have shown that the addition of diethylpyrocarbonate (DEPC) inhibits the binding of Spa to rabbit anti-PVL used as the capturing antibody in ELISA. In Western blotting, the presence of DEPC prevented the binding of detecting antibody to Spa. These modified ELISA and Western blot techniques should prove useful for detecting and quantifying proteins in S. aureus culture supernatants. 2010 Elsevier B.V. All rights reserved.

Citation

Hien M Nguyen, Miguel A Rocha, Koteswara R Chintalacharuvu, David O Beenhouwer. Detection and quantification of Panton-Valentine leukocidin in Staphylococcus aureus cultures by ELISA and Western blotting: diethylpyrocarbonate inhibits binding of protein A to IgG. Journal of immunological methods. 2010 Apr 30;356(1-2):1-5