The use of Endopep-MS for the detection of botulinum toxins A, B, E, and F in serum and stool samples.
Botulinum neurotoxin (BoNT) causes the disease botulism, which can be lethal if untreated. Previous work in our laboratory focused on developing Endopep-MS, a mass spectrometric-based endopeptidase method for the detection and differentiation of BoNT serotypes. We have expanded this effort to include an antibody capture method to partially purify and concentrate BoNT from serum and stool extract samples for the Endopep-MS assay. Because complex matrices such as serum and stool contain abundant endogenous proteases, this technique was needed to remove most proteases from the sample while concentrating BoNT from a sample size of 100 to 500 microl to 20 microl. When this antibody capture method is combined with the Endopep-MS reaction, limits of detection in 500mul of spiked human serum are 10 mouse LD50 (20 mouse LD50/ml) for BoNT A, 0.5 mouse LD50 (1 mouse LD50/ml) for BoNT B, 0.1 mouse LD50 (0.2 mouse LD50/ml) for BoNT E, and 0.5 mouse LD50 (1 mouse LD50/ml) for BoNT F. The limits of detection in spiked stool extracts are somewhat higher due to the high-protease environment of stool extract that also requires use of protease inhibitors. The entire method can be performed in as short a time as 4 h.
Kalb
Suzanne R
SR
Centers for Disease Control and Prevention, National Center for Environmental Health/Agency for Toxic Substances and Disease Registry, Atlanta, GA 30341, USA.
Moura
Hercules
H
Boyer
Anne E
AE
McWilliams
Lisa G
LG
Pirkle
James L
JL
Barr
John R
JR
D000595:Amino Acid Sequence; D000818:Animals; D001905:Botulinum Toxins / Q000032:analysis* / Q000097:blood / Q000737:chemistry; D005243:Feces / Q000737:chemistry*; D013058:Mass Spectrometry / Q000379:methods*; D051379:Mice; D008969:Molecular Sequence Data; D012015:Reference Standards; D012680:Sensitivity and Specificity
D001905:Botulinum Toxins