title: 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.affiliation: National Center for Environmental Health, Centers for Disease Control and Prevention, Atlanta, Georgia 30341, United States.
authors.forename: Zsuzsanna
authors.initials: Z
authors.lastname: Kuklenyik
authors.forename: Anne E
authors.initials: AE
authors.lastname: Boyer
authors.forename: Renato
authors.initials: R
authors.lastname: Lins
authors.forename: Conrad P
authors.initials: CP
authors.lastname: Quinn
authors.forename: Maribel
authors.initials: M
authors.lastname: Gallegos-Candela
authors.forename: Adrian
authors.initials: A
authors.lastname: Woolfitt
authors.forename: James L
authors.initials: JL
authors.lastname: Pirkle
authors.forename: John R
authors.initials: JR
authors.lastname: Barr
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000942:Antigens, Bacterial / Q000737:chemistry*; D001427:Bacterial Toxins / Q000737:chemistry*; D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D010450:Endopeptidases / Q000378:metabolism*; D008969:Molecular Sequence Data; D011817:Rabbits; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D000942:Antigens, Bacterial; D001427:Bacterial Toxins; C030325:anthrax toxin; D010450:Endopeptidases