Rapid Baseline Free Organophosphate Exposure Diagnostic Test

AbstractThe prolific worldwide misuse of organophosphateOPpesticidescoupled with a grave homeland security concern that OP based nerve agents may be deployed either against our troops or civilians on our home soilhas stimulated the search for rapidminiaturized and inexpensive point of carePOCdiagnostic devices for first responders and medical personnel to detect the degree of OP exposureIdentification of the OP agent during an exposure is challenging as it relies on acid hydrolysis products in urine that are rapidly excretedIn contrastacetylcholinesteraseAChEand butylcholinesteraseBChEenzyme activity are established biomarkers for risk assessment because they substantiate exposureas OPs bind directly to AChE and BChEdirectly provide a quantitative biochemical effect of the exposurei eenzyme inhibitionand several weeks post exposureMeasurement of cholinesteraseChEis complicated by interand intraindividual variability of ChE activity and relationship to clinical signs and symptomsThe variability of BChE activity between individuals based simply on geneticssexraceor age is estimated as high asdeviation of the meanThe variation in BChE levels in the population underscores the importance of having individual baseline values of ChE activity to interpret results when assessing exposures to BChE inhibitorsIn the absence of a baselinewhich is likely the case in the event of a mass casualty scenarioBChE inhibition measurements are often based on a population averagewhich introduces a large degree of uncertainty in the reported resultThese uncertainties reduce the sensitivity of BChE activity as a biomarker and thus may provide ambiguous results for subclinicallow dose OP exposureandltenzyme inhibitionIt is critical to note that low cholinesterase inhibition may be subclinical in appearance onlyas there is evidence that even a low dose exposure of OP neurotoxicants can cause long termsignificant chronic neurological deficitsNumerous biochemical methods have been developed to measure ChE levelsincluding the Ellman assayfluorescence assayselectrochemical assaysthe MichelpHChE assayradioactive assaysand the Walter Reed Army Institute of ResearchWRAIRassayHoweverall of these methods are based on detecting a meaningful decrease in enzyme activity from a previously established baselineRecent advances in the application of nanomaterials as transducersrecognition agents or labelsand advances in immunoassay techniques are now being exploited in the design of improved devices for the biomedical detection of non metabolized OP pesticides and nerve agentsmetabolitesand protein adductsOur primary objective is to develop a baseline freebroad spectrum for all OPsfield deployable point of carePOCdevice that confirms and quantitates the degree of exposure to OP neurotoxicants by measuring a ratio of OP bound to unbound BChE Project NarrativeThe overall objective of the work proposed herein is to complete development and preliminary optimization of a prototype point of care device that semi quantitates a ratio of OP adducted BChE to free BChE from human plasma containing titrations of OP to produce a baseline free readoutCompletion of this project will significantly advance the development of a sensitiverapidbaseline freeOP broad spectrumruggedfield deployable point of carePOCdevice that quantitates the degree of exposure to OP neurotoxicants by measuring a ratio of OP bound to unbound BChE as a diagnostic tool for medical personnel administering antidotal therapy