The surface identified by ELISA testing to provide the best combination of sensitivity and signal-to-noise-ratio was fabricated into a SH-SAW device and challenged with a range of Bcl-2 concentrations and another protein to assess sensitivity, selectivity, and performance in a prototype diagnostic sensor. concentration was quantified to demonstrate the effectiveness in surface acoustic wave (SAW)-based detection applications. The surface functionalization approaches exhibited here to specifically and sensitively detect Bcl-2 in a working ultrasonic MEMS biosensor prototype can easily be altered to detect additional biomarkers and enhance other acoustic biosensors. Keywords: bioconjugation, microelectromechanical systems (MEMS), point-of-care, sensor, early detection, ovarian cancer, Bcl-2, surface acoustic wave (SAW), self-assembled monolayer (SAM), polyethylene glycol (PEG) 1.?Introduction Acoustic sensors are capable of measuring physical, chemical and biological quantities using different modes of acoustic (or elastic) waves in various designs and sensor types Gingerol [1]. They have been investigated and used extensively since the 1970s with the introduction of quartz crystal microbalance (QCM) with a selective adsorptive film around the crystal for chemical sensing [2]. Since then, acoustic sensor technology has been Gingerol improved and widely used with the advancements in micro-fabrication technologies, enabling high frequency operation (MHz range) with high sensitivity. Acoustic sensors are typically used as delay line devices or resonators, usually along with electrical components. The typical measurement parameters for sensing include, but are not limited to: insertion loss, phase shift, oscillation frequency, quality factor and impedance [1]. Sensing of different measurands is usually accomplished by applied coatings or thin films that are sensitive to target quantity. The selection of these parameters, quantities and the acoustic mode are affected by the sensor type and design. The most typical acoustic sensor types and related acoustic modes are: surface acoustic wave (SAW) sensors (surface acoustic waves), thickness shear-mode (TSM) sensor (resonant thickness shear modes), shear horizontal acoustic plate mode (SH-APM) sensors (bulk shear horizontal waves), and flexural plate-wave (FPW) sensors (lamb waves) [1]. Each sensor type has its advantages and disadvantages depending on the application for optimal operation and sensitivity. Ovarian cancer is the fifth leading cause of death Tnf among women in the United States and the disease has a 1 in 71 lifetime risk [3]. Reduced lethality is associated with diagnosis in earlier stages of the disease progression [3]. B-cell lymphoma 2 protein (Bcl-2) is currently under investigation as a reliable biomarker for ovarian cancer, and it has been shown that urinary Bcl-2 levels are reliably elevated during different stages of ovarian cancer [4,5]. Based on enzyme-linked immunosorbent assay (ELISA) assessments using urine samples, the average urinary level of Bcl-2 was found to be 0.59 ng/mL in healthy patients, 1.12 ng/mL in benign disorders, 2.60 ng/mL in early-stage ovarian cancer and 3.58 ng/mL in late-stage ovarian cancer [4]. Based on the reliability of urinary levels of Bcl-2 as a biomarker for detecting ovarian cancer at early stages and distinguishing cancer from other gynecological conditions [4], the development of an ultrasonic biosensor has been undertaken to ultimately be used for point-of-care diagnosis. Toward this objective, the device must be able to quantify the biomarker with high sensitivity with minimal false positive results. The biosensor under development employs shear horizontal surface acoustic waves (SH-SAW) in a delay path configuration for their high sensitivity to surface mass loading and the ability to work under liquid loading [6]. It is comprised of a pair of interdigital transducers (IDTs) microfabricated on ST-cut Quartz wafers in the direction 90 off x-axis. The high sensitivity is achieved due to the high SAW velocity of SH waves and the concentration of the wave energy in the surface. A delay path configuration enables surface modifications to a relatively large surface (compared with micro-size scale sensors) to sense the target quantity. The sensing of Bcl-2 binding in the delay path is achieved by monitoring the oscillation frequency change (or shift) of an oscillatory Gingerol circuit, in which the sensor is used as the feedback element. In this sensing method, the oscillation frequency is.