Development of an Epi-Illuminating Fiber Optic Microprobe for Analysis of Chloroplast Specialization and Measurement of Chemical Parameters within Plant Tissue.

本工作将研制一种落射照明光纤微探针 可以用来分析微观环境下的代谢活动 生物组织内的位点。 单色光从一个 可调谐染料激光器将耦合到光纤中， 引导到相对端，在那里有显微尖端（5 μ m 直径），其可以插入生物靶组织内。 光将被用来激发内在的荧光 分子、人造荧光染料或监测吸光度 电子载体和代谢中间产物的变化。 我们 建议在工厂内开发和测试仪器，但在 原理，落射照明探头可以在广泛的应用 各种生物系统。 具体的发展努力将针对（1） 不同吸光度变化的专属性分析 光合电子载体;（2）纤维发育 光学化学传感器，可用于监测代谢 变化，例如细胞和组织内的CO2浓度。 这将通过将聚合物膜与嵌入的 荧光染料到探针尖端， 荧光响应于探针环境中的变化。 落射照明光纤探头将提供信息 在活体内和在活组织内的真实的时间。 的 响应时间快（ns），并且探针将具有高空间分辨率。 分辨率，以便能够测量事件， 光散射组织、细胞内的微位点， 少数（3-4）的生理特化 细胞器如叶绿体。

This work will develop an epi-illumination fiber optic microprobe that could be used to analyze metabolic events at microscopic sites within biological tissues. Monochromatic light from a tunable dye laser will be coupled into an optical fiber and guided to the opposite end where there is a microscopic tip (5 um diameter) that can be inserted within a biological target tissue. The light will be used to stimulate fluorescence of intrinsic molecules, artificial fluorochromes or to monitor absorbance changes of electron carriers and metabolic intermediates. We propose to develop and test the instrument within plants but in principle, the epi-illumination probe could be used in a wide variety of biological systems. Specific developmental efforts will be directed toward (1) analysis of the specialization of different absorbance changes of photosynthetic electron carriers; (2) the development of fiber optic chemical senors that can be used to monitor metabolic changes, such as CO2 concentration, within cells and tissues. This will be pursued by attaching polymeric films with embedded fluorochromes to the probe tip that undergo changes in fluorescence in response to changes in the probe environment. The epi-illumination fiber optic probe will provide information both in vivo and in real time within living tissues. The response time is rapid (ns) and the probe will have high spatial resolution so that it will be possible to measure events at microsites within light-scattering tissues, cells and to examine the physiological specialization of small numbers (3-4) of organelles such as chloroplasts.