IMPROVEMENTS IN FLOW CYTOMETRY OF BACTERIA & VIRUSES

提高流式细胞术灵敏度的几种途径 小细菌和大病毒颗粒的检测与表征 将进行探索，包括一个浸水的流动室，一个激光多普勒 光散射探测器系统、结合光源的前端电子设备 噪声补偿，以及通过声光调制降低光噪声。 作为这些研究的结果，开发了一种高灵敏度的流式细胞仪 将用于光散射和荧光形式的测量 细菌、病毒和直径小于0.5微米的测试颗粒。 光散射和菁染料荧光的双参数测量 在一个样本的多个等量上将被用来估计细菌 并快速确定几种能量中的哪一种 来源将支持一种有机体的生长，以及几种 抑制剂会阻碍增长。比率测量法测量的 荧光素衍生物BCECF的荧光将用于检测pH 细菌细胞质和培养基间的梯度。多站， 光散射和荧光的多参数测量 混合染料，包括膜电位和pH探针，荧光 蛋白质、DNA和RNA以及荧光酶底物的染色将是 调查以确定哪些参数增加了最多的 与细菌鉴定相关的信息。

Several approaches to increasing the sensitivity of flow cytometry for detection and characterization of small bacteria and large virus particles will be explored, including a water-immersed flow chamber, a laser Doppler light scatter detector system, front end electronics incorporating source noise compensation, and light noise reduction by acousto-optic modulation. A high-sensitivity flow cytometer developed as a result of these studies will be used for measurements of light scattering and fluorescence form bacteria, viruses, and test particles smaller than 0.5 mum in diameter. Two-parameter measurements of light scattering and cyanine dye fluorescence on multiple aliquots of a sample will be used to estimate bacterial membrane potentials and to rapidly determine which of several energy sources would support the growth of an organism and which of several inhibitors would prevent growth. Ratiometric measurements of the fluorescence of the fluorescein derivative BCECF will be used to detect pH gradients between bacterial cytoplasm and the medium. Multistation, multiparameter measurements of light scattering and of the fluorescence of mixtures of dyes, including membrane potential and pH probes, fluorescent stains for protein, DNA and RNA, and fluorogenic enzyme substrates, will be investigated to determine which parameters add the most additional information relevant to bacterial identification.