Automatic Assay of Blood Cholinesterase in the Field

There is a critical need for rapid, accurate measurement of serum cholinesterase activity in the field to provide a timely determination of nerve agent exposure. The overall objective of the project is to develop an easy-to-use analytical system consistingof small, inexpensive (and therefore disposable) silicon microchips used with a portable instrument, thus permitting field cholinesterase measurements by unskilled personnel. Phase I demonstrated the feasibility of adapting silicon microsampling and assaydevices, originally developed for blood glucose monitoring, to cholinesterase measurement. These microchips have integrated microneedles capable of penetrating the skin and taking a blood sample painlessly and reliably. An electrochemical redox-mediatedcholinesterase assay was developed (providing a potentially long shelf-life) and silicon microchips with novel microfluidics were designed and fabricated. In Phase II, the on-chip cholinesterase assay and microfluidics will be optimized, the completemicrochip with silicon microneedle packaged, a prototype instrument designed and built, and the integrated system evaluated. The proposed instrument with sophisticated microfluidics and innovative electrochemical readout will measure enzymatic activity invery small blood samples (as small as 2 nanoliters). The blood sample, drawn by capillary force into the chip, flows through a microchannel to react with a reagent solution containing a substrate to the enzyme. Electrochemical sensors with specificity fora reaction product measure product concentrations and provide a readout indicative of cholinesterase activity in the blood sample. This unique one-step automatic painless blood testing system will enable rapid and accurate measurement of cholinesterase activity in the field for evaluation of exposure to nerve agents in both military and terrorism contexts. It will also provideeasy, reliable monitoring of chemical and agricultural workers for toxic pesticide exposure. With the addition of other analytes, this platform technology is widely applicable to other military and civilian blood testing market segments.