High-Throughput Immunomagnetic Scavenging Technique for Quantitative Analysis of Live VX Nerve Agent in Water, Hamburger, and Soil Matrixes

Abstract

We have developed a novel immunomagnetic scavenging technique for extracting cholinesterase inhibitors from aqueous matrixes using biological targeting and antibody-based extraction. The technique was characterized using the organophosphorus nerve agent VX. The limit of detection for VX in high-performance liquid chromatography (HPLC)-grade water, defined as the lowest calibrator concentration, was 25 pg/mL in a small, 500 μL sample. The method was characterized over the course of 22 sample sets containing calibrators, blanks, and quality control samples. Method precision, expressed as the mean relative standard deviation, was less than 9.2% for all calibrators. Quality control sample accuracy was 102% and 100% of the mean for VX spiked into HPLC-grade water at concentrations of 2.0 and 0.25 ng/mL, respectively. This method successfully was applied to aqueous extracts from soil, hamburger, and finished tap water spiked with VX. Recovery was 65%, 81%, and 100% from these matrixes, respectively. Biologically based extractions of organophosphorus compounds represent a new technique for sample extraction that provides an increase in extraction specificity and sensitivity.

Development of immunosorbents coupled on-line to immobilized pepsin reactor and micro liquid chromatography–tandem mass spectrometry for analysis of butyrylcholinesterase in human plasma

“…Hence, it is essential to extract HuBuChE from highly concentrated proteins in plasma before digestion and analysis by LC/MS to avoid ion suppression. Current purification methods are based on HuBuChE extraction from plasma by affinity chromatography on procainamide gels [4,8,21,22] or by antibodies immobilized on magnetic beads activated with streptavidin or protein G [23][24][25][26][27]. Procainamide gels are relatively cheap but not selective enough, implying the need of additional purification steps, which extends the extraction time and reduces the total recovery of the method [4,28].…”

“…Purification of HuBuChE by immunomagnetic beads is more specific than the procainamide gels approach and requires smaller amounts of plasma. In most protocols, HuBuChE is directly digested on the immunomagnetic beads in contact with the protease, which involves the use of a new batch of antibodies for each sample since they are also digested [12,13,[25][26][27]. The digestion step leads to peptides derived from HuBuChE but also from antibodies, protein G, protease by self-digestion and thus to numerous peptides that could affect the mass spectrometry response of the target peptide adducts.…”

“…In addition, IS can be directly coupled on-line to digestion on IMER and to LC/MS analysis, which avoids sample loss and contamination and allows automation of the whole analytical procedure. Among different grafting supports available for antibody immobilization, like silica [30,31], polystyrene divinyl benzene [32], glycidyl methacrylate-coethylene glycol dimetacrylate (GMA-EDTA) [33,34] or magnetic beads [13,25,26,35], two supports, i.e. epoxy activated polymethacrylate resin [36] and CNBr activated sepharose [19,37] were chosen.…”

Online coupling of immunoextraction, digestion, and microliquid chromatography-tandem mass spectrometry for the analysis of sarin and soman-butyrylcholinesterase adducts in human plasma

“…Since HuBuChE is ten times more abundant than AChE, located in plasma and more sensitive to inhibition by some OPNAs than AChE, analysis of OPNA-HuBuChE adducts has been preferred [20][21][22]. In most of the studies, HuBuChE is purified and digested by pepsin into the nonapeptide 3 195 FGESAGAAS 203 which includes the OPNA moiety fixed on the 198 serine residue and the adducted nonapeptide is further identified by LC/MS 2 [7,18,23,24]. After a few minutes for soman to a couple of days for VX, the alkoxy-group of the adducted OPNA moiety is cleaved and replaced by a hydroxyl function during a phenomenon called ageing [25].…”

Extraction and Analysis of V-Type Agents (VX, RVX, CVX, and VM) from Various Food Matrices by Ultra-Performance Liquid Chromatography-Time-Of-Flight Mass Spectrometry

“…10,11 The pesticide literature often includes sample-preparation techniques that are commercially available and affordable, such as solid-phase extraction cartridges [12][13][14][15][16][17][18][19][20] or QuEChERS systems (Quick, Easy, Cheap, Effective, Rugged, and Safe); [21][22][23][24][25][26][27][28][29] however, the CWA literature seems to focus more on new techniques and specialized equipment that may not be as readily accessible to every laboratory. 9,[30][31][32][33][34][35] This document reports the efforts of the Agent Chemistry Team from the Research and Technology Directorate of the U.S. Army Edgewood Chemical Biological Center (ECBC; Aberdeen Proving Ground, MD) in developing new extraction and analytical detection methodologies using liquid chromatography-mass spectrometry (LC-MS). The objective of this task was to provide development and laboratory support for extraction of V-type agents ( Figure 1) from various food samples.…”