Detection and quantification of anthrax lethal factor in serum by mass spectrometry.
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| Abstract |
The lethal toxin produced during Bacillus anthracis infection is a complex of protective antigen, which localizes the toxin to the cell receptor, and lethal factor (LF), a zinc-dependent endoproteinase whose known targets include five members of the mitogen-activated protein kinase kinase (MAPKK) family of response regulators. We have developed a method for detecting functional LF in serum. Anti-LF murine monoclonal antibodies immobilized on magnetic protein G beads were used to capture and concentrate the LF from serum. The captured LF was exposed to an optimized MAPKK-based peptide substrate, which it hydrolyzed into two smaller peptides. The LF cleavage products were then analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MS) and quantified by isotope dilution-MS. The entire analytical method can be performed in less than 4 h with detection of LF levels as low as 0.05 ng/mL. The method was used to quantify LF levels in serum from rhesus macaques infected with B. anthracis. Serum samples obtained at day 2 postinfection contained 30-250 ng/mL LF and illustrated the clear potential to detect LF earlier in the infection cycle. This method represents a highly specific and rapid diagnostic tool for early anthrax and has a potential additional role as a research tool for understanding toxemia and effects of medical countermeasures for anthrax.
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| Authors | Anne E Boyer, Conrad P Quinn, Adrian R Woolfitt, James L Pirkle, Lisa G McWilliams, Karen L Stamey, Dennis A Bagarozzi, John C Hart Jr, John R Barr |
| Journal | Analytical chemistry (Anal Chem) Vol. 79 Issue 22 Pg. 8463-70 (Nov 15 2007) ISSN: 0003-2700 [Print] United States |
| PMID | 17929949 (Publication Type: Journal Article) |
| Chemical References |
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