König K
Institute of Anatomy II, Friedrich Schiller University, Jena, Germany.
Cell Mol Biol (Noisy-le-grand). 1998 Jul;44(5):721-33.
The most important application of continuous wave (cw) near infrared (NIR) microbeams in cellular and molecular biology are single-beam gradient force optical traps, also called "laser tweezers". Laser tweezers have been used for optical picoNewton force determination as well as for 3D cellular and intracellular micromanipulation, such as optical spermatozoa transportation in laser-assisted in vitro fertilization. Light intensities in the MW/cm2 range are necessary to confine motile spermatozoa in the optical trap. The enormous photon concentration in space and time results in non-resonant two-photon excitation of endogenous and exogenous absorbers with electronic transitions in the ultraviolet and visible spectral range. Trap-induced two-photon excitation of intracellular flurorophores can be used to study metabolism and vitality of motile cells without additional fluorescence excitation sources. Therefore, laser tweezers as sources of two-photon excitation may act as novel non-linear tools in cell diagnostics. The far red/NIR trapping radiation, in particular <800 nm, may also excite endogenous absorbers such as NAD(P)H, flavins, porphyrins and cytochromes. Excitation of these cellular absorbers may result in oxidative stress via type I and type II photooxidation processes. Severe non-linear-induced cell damage in a variety of cells confined in <800 nm traps was found. Two-photon induced destructive effects are enhanced in multimode traps due to longitudinal mode-beating phenomena. Pulsed laser sources are not suitable for safe optical trapping of living cells. The use of single frequency long-wavelength NIR traps (800 nm-1200 nm) for vital cell handling is recommended.
连续波（cw）近红外（NIR）微光束在细胞和分子生物学中最重要的应用是单光束梯度力光阱，也称为“激光镊子”。激光镊子已被用于光学皮牛顿力的测定以及三维细胞和细胞内的微操作，如在激光辅助体外受精中进行光学精子运输。在光阱中捕获活动精子需要MW/cm²范围内的光强。在空间和时间上极高的光子浓度会导致内源性和外源性吸收体在紫外和可见光谱范围内发生非共振双光子激发。陷阱诱导的细胞内荧光团双光子激发可用于研究活动细胞的代谢和活力，而无需额外的荧光激发源。因此，作为双光子激发源的激光镊子可能成为细胞诊断中的新型非线性工具。远红/NIR捕获辐射，特别是<800 nm的辐射，也可能激发内源性吸收体，如NAD(P)H、黄素、卟啉和细胞色素。这些细胞吸收体的激发可能通过I型和II型光氧化过程导致氧化应激。发现在<800 nm陷阱中捕获的各种细胞中存在严重的非线性诱导细胞损伤。由于纵向模式拍频现象，多模陷阱中双光子诱导的破坏效应会增强。脉冲激光源不适用于活细胞的安全光学捕获。建议使用单频长波长NIR陷阱（800 nm - 1200 nm）进行活细胞处理。
