Physiological monitoring of optically trapped cells: assessing the effects of confinement by 1064-nm laser tweezers using microfluorometry

Abstract

We report the results of microfluorometric measurements of physiological changes in optically trapped immotile Chinese hamster ovary cells (CHOs) and motile human sperm cells under continuous-wave (CW) and pulsed-mode trapping conditions at 1064 nm. The fluorescence spectra derived from the exogenous fluorescent probes laurdan, acridine orange, propidium iodide, and Snarf are used to assess the effects of optical confinement with respect to temperature, DNA structure, cell viability, and intracellular pH, respectively. In the latter three cases, fluorescence is excited via a two-photon process, using a CW laser trap as the fluorescence excitation source. An average temperature increase of < 0.1 +/- 0.30 degrees C/100 mW is measured for cells when held stationary with CW optical tweezers at powers of up to 400 mW. The same trapping conditions do not appear to alter DNA structure or cellular pH. In contrast, a pulsed 1064-nm laser trap (100-ns pulses at 40 microJ/pulse and average power of 40 mW) produced significant fluorescence spectral alterations in acridine orange, perhaps because of thermally induced DNA structural changes or laser-induced multiphoton processes. The techniques and results presented herein demonstrate the ability to perform in situ monitoring of cellular physiology during CW and pulsed laser trapping, and should prove useful in studying mechanisms by which optical tweezers and microbeams perturb metabolic function and cellular viability.

Optical tweezers for single cells

“…When using 100 mW laser tweezers (1064 nm in wavelength), the trap-induced temperature increase is between 1 and 28C. In trapped human sperm cells, hamster ovary cells and liposomes, the temperature increases were measured to be of the order of 10, 11.5 and 14.58C W K1 , respectively (Liu et al 1995(Liu et al , 1996. This is confirmed by Peterman et al (2003).…”

Optical Trapping of Nanoparticles

“…Trapping of biological materials of large polarizability and size has included bacteria 3 , living cells 17,2,18 , the tobacco mosaic virus 3 and manipulation and stretching of DNA strands tethered at the ends with large dielectric particles 19 ; however, direct trapping of smaller biological samples without tethering remains challenging. This trapping configuration is capable of trapping small dielectric particles at lower light intensities than conventional light tweezers and the circular nanohole, allowing small biological particles to be held for long periods of time without damage or tethering.…”

Exploitation of physical and chemical constraints for three-dimensional microtissue construction in microfluidics

“…The temperature increases as much as 10, 11.5, and 14.5°C / W, respectively. 139,140 Local heating is a serious issue in trapping cells as heat adversely affects enzyme activity and other sensitive cellular functions; also, steep thermal gradient might cause a convection current that disrupts laminar flow in the microfluidic channel. Therefore, there is much room to improve and potentially in combination with other physical and chemical trapping techniques.…”