(388e) Development of a Unique Dual Ionophore Ion Selective Electrode for the Detection of Proteins and Cells

The need for novel, rapid, and inexpensive biosensor platforms is continuously increasing as the demand for point-of-care capable technology grows. Diagnostic tools which provide results within 30 minutes outside of a central laboratory setting are especially critical for the detection of rapidly progressing, severe medical conditions such as sepsis. Sepsis, which occurs due to a systemic inflammatory immune response to bacterial, viral, or fungal infections, is a major cause of death in the United States. Novel biosensors which allow point-of-care, rapid identification of pathogenic species could dramatically improve patient survival odds by allowing more timely administration of targeted antibiotic therapy. Potentiometric-based biosensors such as ion selective electrodes (ISEs) provide a promising platform for point-of-care detection, as they boast response times on the order of seconds, an electronic signal which can be easily recorded on a simple hand-held electronic device, and inexpensive construction. In this presentation, we will describe the design of a unique ISE modeled after electrochemical potential regulation in living cells and neurons. The dual ionophore ISE (dI-ISE) incorporates two separated ion selective membranes, one sensitive to potassium and the other to sodium. The two membranes are electrically connected by identical inner and outer electrolyte solutions; thus, the equilibrium electric potential developed across the sensor is dependent on the concentration of both ion species near the membrane surfaces, as occurs in living cells. The dI-ISE can be adapted for biosensing applications by the covalent attachment of a biorecognition molecule to the surface of one of the ion selective membranes. Upon analyte binding, the surface of the membrane is partially covered, and resistance to ion transport near the surface of the membrane increases. This causes a shift in the measured potential, such as occurs in living cells or neurons upon closure of selective ion channels. Through the incorporation of the second membrane and ion species, a stable electric potential still exists in the system even when the transport of one ion is completely blocked, allowing a true shift in potential to be measured. Results from studies which simulated protein or cell binding using inert materials such as rubber covers or silicone glue to control membrane surface coverage will be presented. Additionally, a mathematical model based on a modified form of the Goldman-Hodgkin-Katz equation will be presented to describe the surface-coverage based response of the dI-ISE system. Finally, preliminary results from potentiometric and microscopy studies with biotin-functionalized dI-ISE membranes will be presented to show the potential of the system to be used for biosensing applications. These initial studies show significant streptavidin capture on the membrane surface, visualized through fluorescent microscopy, and small shifts in potential after short 30 minute incubations with streptavidin solutions. Preliminary results clearly demonstrate the potential of the dI-ISE for further biosensing applications, including the detection of bacterial and viral species and rare proteins or cells.