Time-resolved imaging was used to examine the use of pulsed laser microbeam irradiation to produce cell lysis. Lysis was accomplished through the delivery of 6 ns, lambda=532 nm laser pulses via a 40x, 0.8 NA objective to a location 10 microm above confluent monolayers of PtK2 cells. The process dynamics were examined at cell surface densities of 600 and 1000 cells/mm2 and pulse energies corresponding to 0.7x, 1x, 2x, and 3x the threshold for plasma formation. The cell lysis process was imaged at times of 0.5 ns to 50 micros after laser pulse delivery and revealed the processes of plasma formation, pressure wave propagation, and cavitation bubble dynamics. Cavitation bubble expansion was the primary agent of cell lysis with the zone of lysed cells fully established within 600 ns of laser pulse delivery. The spatial extent of cell lysis increased with pulse energy but decreased with cell surface density. Hydrodynamic analysis indicated that cells subject to transient shear stresses in excess of a critical value were lysed while cells exposed to lower shear stresses remained adherent and viable. This critical shear stress is independent of laser pulse energy and varied from approximately 60-85 kPa for cell monolayers cultured at a density of 600 cells/mm2 to approximately 180-220 kPa for a surface density of 1000 cells/mm2. The implications for single cell lysis and microsurgery are discussed.

Water under tension, as can be found in several systems including tree vessels, is metastable. Cavitation can spontaneously occur, nucleating a bubble. We investigate the dynamics of spontaneous or triggered cavitation inside water filled microcavities of a hydrogel. Results show that a stable bubble is created in only a microsecond timescale, after transient oscillations. Then, a diffusion driven expansion leads to filling of the cavity. Analysis reveals that the nucleation of a bubble releases a tension of several tens of MPa, and a simple model captures the different time scales of the expansion process.

We use time-resolved imaging to examine the lysis dynamics of non-adherent BAF-3 cells within a microfluidic channel produced by the delivery of single highly-focused 540 ps duration laser pulses at λ = 532 nm. Time-resolved bright-field images reveal that the delivery of the pulsed laser microbeam results in the formation of a laser-induced plasma followed by shock wave emission and cavitation bubble formation. The confinement offered by the microfluidic channel constrains substantially the cavitation bubble expansion and results in significant deformation of the PDMS channel walls. To examine the cell lysis and dispersal of the cellular contents, we acquire timeresolved fluorescence images of the process in which the cells were loaded with a fluorescent dye. These fluorescence images reveal cell lysis to occur on the nanosecond to microsecond time scale by the plasma formation and cavitation bubble dynamics. Moreover, the time-resolved fluorescence images show that while the cellular contents are dispersed by the expansion of the laser-induced cavitation bubble, the flow associated with the bubble collapse subsequently re-localizes the cellular contents to a small region. This capacity of pulsed laser microbeam irradiation to achieve rapid cell lysis in microfluidic channels with minimal dilution of the cellular contents has important implications for their use in lab-on-a-chip applications.