| Laser-induced optical breakdown (LIOB) is a nonlinear energy absorption process that can generate precise damage in biological tissues. With femtosecond laser pulses, disruption is highly localized with minimal thermal and mechanical effects to the surrounding region. Cavitation bubbles are produced as a result of LIOB, and these bubbles can be detected and monitored with high-frequency ultrasound. In this work, the controllable viability effects of LIOB bubbles in single cells were characterized.;Using a high-frequency acoustic transducer synchronized with a 793 nm, 100 fs laser pulsed at 250 kHz, thermal effects in the vicinity of an LIOB event were directly assessed. Temperaturedependent pulse-echo displacements were calculated using phase-sensitive correlation tracking and fit to a finite-element heat transfer model to estimate thermal distribution. Results indicate a minimal temperature increase (<1 degree C) within 100 microns of a bubble created with multiple laser pulses, confirming that LIOB can be controlled to be thermally noninvasive in the bubble vicinity.;Acoustically detectable microbubbles were generated in individual cells with femtosecond LIOB. By adjusting laser fluence, exposure time, and focal location, LIOB could be controlled to produce distinctly different cellular effects. Small (1-2 micron) bubbles with short lifetimes (10100 ms) could be generated in cells without affecting their viability; and, alternatively, large (510 micron) bubbles with long lifetimes (1-5 s) could be generated for selective cell killing without affecting immediately neighboring cells. Experiments were performed in Chinese hamster ovary (CHO) cells in vitro, and LIOB was detected with both optical and acoustic microscopy. A long-term proliferation assay was also performed using green-fluorescent MCA207 mouse sarcoma cells targeted for LIOB. This assay confirmed that nondestructive bubbles did not affect target cell proliferation over several generations, and that destructive bubbles could indeed eliminate target cells and prevent further proliferation with no effect on immediately neighboring cells.;These studies help to outline future applications for site-activated, acoustically monitored intracellular microbubbles. Nondestructive bubbles can potentially be used for functional cell measurements without introducing exogenous agents or affecting subsequent cell proliferation, and destructive bubbles can be used for highly precise biologically-targeted cancer cell therapy with real-time acoustic validation. |
