Comparison of MALDI-TOF-MS and HPLC-ESI-MS/MS for endopeptidase activity-based quantification of Anthrax lethal factor in serum.

Abstract

Diagnosing and treating anthrax at the earliest stage of disease is critical. We developed a method to diagnose anthrax at early stages of infection by detecting anthrax lethal factor (LF) at the attomol/mL level in plasma or serum. This method uses antibody capture and quantification of LF endoproteinase activity by isotope dilution matrix-assisted laser-desorption ionization (MALDI) time-of-flight (TOF) mass spectrometry (MS). Many public health laboratories do not use MALDI-TOF-MS; thus, we have adapted the LF method for detection by electrospray ionization (ESI) tandem MS (MS/MS), which allowed comparison of both MS platforms for LF quantification. Calibration curves were linear from 0.05-2.5 ng/mL when measured after 2 h and from 0.005-1.0 ng/mL after 18 h incubation time. The limit of detection was 0.005 ng/mL using a 200 μL sample. The coefficient of variation for quality control samples was 6-12% for both MS platforms. Samples used to perform cross-validation included 158 serum samples from a study in rabbits exposed to anthrax spores by inhalation. Some were treated with anthrax immune globulin before exposure. Concentrations measured by ESI-MS/MS matched those by MALDI-TOF-MS with p = 0.99 (r(2) = 0.997) and -0.25% mean relative difference (±9% standard deviation). This study shows that isotope dilution MALDI-TOF-MS is a robust and precise quantitative MS platform.

• Authors: Zsuzsanna Kuklenyik, Anne E Boyer, Renato Lins, Conrad P Quinn, Maribel Gallegos-Candela, Adrian Woolfitt, James L Pirkle, John R Barr
• Journal: Analytical chemistry (Anal Chem) Vol. 83 Issue 5 Pg. 1760-5 (Mar 01 2011) ISSN: 1520-6882 [Electronic] United States
• PMID: 21302970 (Publication Type: Journal Article)

Chemical References

• Antigens, Bacterial
• Bacterial Toxins
• anthrax toxin
• Endopeptidases

Topics

• Amino Acid Sequence
• Animals
• Antigens, Bacterial (chemistry)
• Bacterial Toxins (chemistry)
• Calibration
• Chromatography, High Pressure Liquid (methods)
• Endopeptidases (metabolism)
• Molecular Sequence Data
• Rabbits
• Spectrometry, Mass, Electrospray Ionization (methods)
• Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization (methods)
• Tandem Mass Spectrometry (methods)