Sonoporation of cultured cells in the rotating tube exposure system

Suspensions of Chinese hamster ovary cells were exposed to ultrasound in th e presence of fluorescent dextran to determine the conditions needed for so noporation with uptake of the large molecules, Albunex(R), a gas-body- base d ultrasound contrast agent, was added to enhance cavitation. Ultrasound wa s continuous wave at frequencies of 1.0, 1.68, 2.25, 3.3, 5.3, and 7.15 MHz , Sterile 4.5-mL exposure chambers were rotated at 60 rpm to promote cavita tion activity during the 1-min exposures. After exposure, cells were tested for sonoporation by counting fluorescent cells and for cell lysis by count ing cells stained by trypan blue. Sonoporation was a sensitive bioeffects i ndicator that was detected at pressure amplitudes lower than were needed fo r transient cavitation or cavitation-induced cell lysis, For 10% Albunex, a pparent thresholds for sonoporation, which were comparable to the levels re quired to perturb the gas bodies, were 0.084 MPa (spatial peak negative pre ssure amplitude) from 1.0-3.3 MHz and 0.27 MPa at 5.3 and 7.15 MHz. Sonopor ation decreased slightly if the tube was not rotated. The effects increased for increasing Albunex concentration (with rotation), The plating efficien cy of cells exposed to 0.2, MPa at 2.25 MHz and sorted by a flow cytometer was 19% (3.6% standard deviation [SD]) for fluorescent cells, compared to 6 7% (1% SD) for nonfluorescent exposed cells and 62% (6% SD) for sham-expose d cells. The reduced viability represents an important consideration for po ssible applications of sonoporation. (C) 1998 World Federation for Ultrasou nd in Medicine & Biology.