Examination of laser microbeam cell lysis in a PDMS microfluidic channel using time-resolved imaging

Abstract

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 lambda = 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 time-resolved 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.

Fig. 1

(a) Top view of microfluidic chip design. (b) Experiment geometry.

Fig. 2

Experiment setup for time-resolved fluorescence imaging.

Fig. 3

Probability of plasma formation within the microfluidic channel as a function of pulse energy. The plasma threshold energy and sharpness governing the probability as predicted by the Gaussian error function are E_p = 2.29 ± 0.08 μJ and S = 1.4 ± 0.3 μJ⁻¹. See text for further details.

Fig. 4

Bright-field images of the cavitation bubble dynamics inside the microfluidic channel on times scales spanning 9 orders of magnitude from 10 ns to 10 s. Scale bar = 50 μm.

Fig. 5

Bright-field images of a BAF-3 cell within the microchannel before and after delivery of the pulsed laser microbeam in the microfluidic channel. Scale bar = 50 μm.

Fig. 6

Fluorescent cell lysis dynamics inside the microfluidic chip. Fluorescent images of the laser-microbeam cell lysis process inside the microfluidic channel on times scales spanning 9 orders of magnitude from 10 ns to 10 s. Scale bar = 50 μm.

Fig. 7

Attempted lysis of the cell not positioned in the center of the microfluidic channel. Scale bar = 50 μm.

Fig. 8

(a) Cell before lysis, (b) after lysis (scale bar = 50 μm). The size of the two bottom frames is 50 × 50 μm.