A novel technique for measuring ultrasound backscatter from single micron-sized objects

The measurement of the ultrasound backscatter from individual micron-sized objects is required to gain an understanding of the behavior of both weak (cells) and strong (contrast agents) scatterers for applications ranging from tissue characterization to molecular imaging. However, obtaining such a response remains a challenge. For instance, the presence of air bubbles in a suspension of cells during measurements of cell backscatter may lead to the incorrect interpretation of the backscattered signals. In addition, the size and shape of the single object that produces an ultrasound backscatter signal are critical input parameters to theoretical models, yet hard to be measured experimentally. In this work, a novel technique combining a Xenoworks microinjection system (Sutter, Inc., Los Angeles, CA) with co-registered Olympus IX71 inverted microscope (Olympus America, Inc., Center Valley, PA) and a VEVO770 Ultrasound imaging device (VisualSonics, Inc., Toronto, ON) was developed in which the ultrasound backscatter response from a single object was obtained under optical microscope guidance. This technique provides accurate information about the size and shape of the object. Two transducers of central frequencies of 25 and 55 MHz were used (for a total spectrum of 12-57 MHz). The foci of the optical lens and the transducer were aligned to obtain optical and ultrasonic images of the same region. The object of interest was attached to the micropipette (using negative pressure) and then released from the micropipette (using positive pressure and/or tapping on the micropipette) while imaging it both optically and ultrasonically. In order to calibrate the system, a micropipette was used to grab a 20 ?m polystyrene microsphere from a suspension of microspheres in degassed water by applying a pressure of -18.9 kPa. The microsphere was released by applying a pressure of +35.0 kPa. During the release, optical and ultrasonic raw RF lines were obtained. These lines were then used to obtain the power spectral plot of individual microspheres which were compared to analytical solutions. A very good agreement was found (error of 1%) between the measured backscatter response of microspheres and that of a Faran model of an elastic sphere. Extension of this method to prostate carcinoma (PC-3) cells showed a good agreement (error of 5%) when compared to the Anderson fluid sphere model. This technique is capable of providing accurate measurements of the backscatter from individual objects and is currently being used to deduce the backscatter response from other cell lines of different sizes and from ultrasound contrast agents either in isolation or when attached to a cell. The advantages of the technique along with its future applications are discussed.

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