Manipulation of in vitro toxicant sensors in an ultrasonic standing wave.

Multi-cellular spheroids are increasingly employed as in vitro sensors of toxicants and a single spheroid can be used as a test object. An ultrasonic standing wave trap (USWT) can hold small particles in a medium-flowing system. This study investigated the conditions for holding HepG2 spheroids in an USWT and its relevance to use in toxicity testing. It can take many hours to reach a detectable end point of cell damage in a standard cellular in vitro toxicant assay and the process might be accelerated through increased sample flow past the spheroid. A USWT was employed here to levitate and hold HepG2 spheroids stationary against a flow of 3 mm s(-1) when the acoustic pressure amplitude is 1.9 MPa. The ultrasonic drive frequency was 1.64 MHz. Acoustic microstreaming in the standing wave chamber generated 1 mm s(-1) flow past a levitated spheroid-scale (80 microm diameter) latex particle in the absence of sample through-flow. The conditions required to form aggregates of cells of a HepG2 cell line in a single half wavelength ultrasonic standing wave mini-chambers are also described here. It is argued that the manipulation capabilities demonstrated may have potential in increasing the efficiency of in vitro toxicant detection by spheroids. Preliminary, visual (unquantified) fluorescence microscopy observations of spheroids levitated in the standing wave in the presence of the toxicant DL-propranolol do suggest accelerated loss of viability compared with controls.