MEASURE OF PLATELET INTRACELLULAR FREE CALCIUM ION CONCENTRATION: CELL BEHAVIOR

比率荧光显微镜已被用于成像和测量 细胞内游离钙浓度，[Ca++]i， 血小板在生物材料基质上附着和扩散。 的 基质的组成和化学性质的特征在于： ESCA。 一种新型的钙离子敏感荧光团Fura Red（tm） 并实现了系统的原位校准。 在静息细胞中观察到[Ca++]i为50- 100 nM。 [Ca++]i 发现对材料接触的反应是不均匀的。 以下 可变的滞后期长达5分钟，响应细胞表现出 [Ca++]i升高100- 300 nM。 其他血小板显示很少或 与表面接触的[Ca++]i无变化。 那些血小板 证明峰[Ca++]i值大于350 nM， 在实验期间传播，而那些没有传播的 观察到[Ca++]i值保持低于250 nM;这表明 血小板扩散与[Ca++]i升高有关。 钙 还对粘附在来自一个细胞的表面上的血小板进行了成像。 流动细胞悬液。 结果发现，血小板附着， 显示伪足形成的早期迹象， [Ca++]i在最初的几分钟内增加， contact. 结果表明，一种新的方法可视化的 接触刺激血小板的信号转导过程 为进一步研究生物材料中[Ca++]i的变化奠定了基础 血小板中的信号传导和[Ca++]i的可能相关性 与血小板活化的其他方面反应，如颗粒 释放和促进血液凝固。

Ratio fluorescence microscopy has been used to image and measure the intracellular free calcium concentration, [Ca++]i, in single platelets as they attach and spread on biomaterial substrates. The composition and chemistry of the substrates were characterized by ESCA. A novel Ca++-sensitive fluorophore, Fura Red(tm) was loaded into platelets and an in situ calibration of the system was achieved. The [Ca++]i in resting cells was observed to be 50-100nM. The [Ca++]i response to material contact was found to be heterogeneous. Following a variable lag period of up to 5 minutes, responding cells exhibited a rise in [Ca++]i of 100-300nM more. Other platelets showed little or no change in [Ca++]i in contact with the surface. Those platelets demonstrating peak [Ca++]i values greater than 350nM were found to have spread during the experiment, while those that did not spread were seen to have [Ca++]i values remaining below 250nM; suggesting that platelet spreading is associated with a rise in [Ca++]i. Calcium imaging was also performed on platelets adhering to surfaces from a flowing cell suspension. It was found that platelets attached and showed early signs of pseudopodia formation without significant increases [Ca++]i during the initial few minutes of contact. The results demonstrate a novel method of visualizing the process of signal transduction in platelets stimulated by contact with biomaterials, and lay the groundwork for further study of [Ca++]i signaling in platelets and of the possible correlation of the [Ca++]i response with other aspects of platelet activation such as granule release and the promotion of blood coagulation.