Spectral evaluation of laser-induced cell damage with photothermal microscopy

Abstract

Determining cell photo-damage is important for laser medicine and laser safety standards. This work evaluated the potential of photothermal (PT) technique for studying invasive laser-cell interaction, with a focus on PT evaluation of spectral dependence of laser-induced damage in visible region at single intact cell level. PT is based on irradiation of a single intact cells with a tunable pump laser pulse (420-570 nm, 8 nanoseconds, 0.1-300 microJ) and monitoring of temperature-dependent variations of the refractive index with a second, collinear probe beam in pulse (imaging) mode (639 nm, 13 nanoseconds, 10 nJ), and continuous (integrated PT response) mode (633 nm, 2 mW). The local and the integrated PT responses from the individual living red blood cells, lymphocytes, and cancer cells (K562) in vitro were obtained at different pump laser fluence and wavelength and compared with data obtained by conventional viability tests (Annexin V--propidium iodide). The cell damage with pump pulse lead to specific change in PT response's temporal shape and PT image's structure. The photodamage thresholds varied in the range of 0.5-5 J/cm2 for red blood cells, 4.4-42 J/cm2 for lymphocytes, and 36-90 J/cm2 for blast cells in the pump wavelength range of 417-555 nm. Damage threshold at different wavelength depends on absorption spectra of cells. Spectral evaluation of laser-damage thresholds can be done in two supplements for each PT mode--PT imaging and integrated PT response. The correlation between specific change of PT parameters and cell damage permits using PT technique to rapidly estimate the invasive conditions of the laser-cell interactions.