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Applications of fixed-time flow cytometry in cell biology,

机译：固定时间流式细胞术在细胞生物学中的应用，

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Abstract: Flow cytometric (FCM) measurement of intracellular free calcium $LB@Ca$+2$PLU$/$RB$-i$/, transients is usually done by two methods: (a) after a short prerun period to assess the baseline the measurement is stopped, stimulus is added and the measurement continued or (b) stimulus is injected during measurement and the sample pressure briefly increased to deliver cells rapidly to the detection point. In (a) measurement of very short transients $LB@Ca$+2$PLU$/$RB$-i$/ is impeded by the lag time between stimulus addition and restart of acquisition. In (b) response of pressure sensitive cells is hard to analyze. Furthermore, (a) and (b) do not allow to quantify and sort rare responders. A simple Fixed- Time device has been developed. Ca$+2$PLU$/ sensitive fluorescent dye labeled cells and a stimulus are placed in different vials. Both fluids are forced by the same pressure through tubing that merges into a T-junction where they mix and are delivered through a connecting tube to the FCM: $LB@Ca$+2$PLU$/$RB$-i$/ is measured at a certain time after stimulation that is adjusted by sample flow rate and length of the connecting tube. With Fixed-Time, the pressure sensitive neuronal NH15-CA2 cell was analyzed. Furthermore, rare neurotransmitter responsive fibroblast from normal and transfected cultures were sorted and cloned and their dose response characterized. The results demonstrate that fixed- time FCM is an important tool for the analysis of the cells physiology and the preparation of responders. !23
机译：摘要：流式细胞术（FCM）测量细胞内游离钙$ LB @ Ca $ + 2 $ PLU $ / $ RB $ -i $ /，瞬态通常通过两种方法完成：（a）经过短暂的运行前评估基线停止测量，添加刺激并继续进行测量，或者（b）在测量过程中注入刺激，短暂增加样品压力以将细胞快速递送至检测点。在（a）中，非常短的瞬态的测量$ LB @ Ca $ + 2 $ PLU $ / $ RB $ -i $ /被刺激添加和重新开始获取之间的延迟时间所阻碍。在（b）中，压敏细胞的响应难以分析。此外，（a）和（b）不允许对稀有响应者进行量化和分类。开发了一种简单的固定时间设备。 Ca + 2 $ PLU $ /敏感的荧光染料标记细胞和刺激物置于不同的小瓶中。两种流体在相同的压力下通过管道汇入T形接头，并在那里混合并通过连接管输送到FCM：$ LB @ Ca $ + 2 $ PLU $ / $ RB $ -i $ /在刺激后的某个时间测量，可通过样品流速和连接管的长度进行调整。使用固定时间分析压敏神经元NH15-CA2细胞。此外，对来自正常和转染培养物的稀有神经递质反应性成纤维细胞进行了分类和克隆，并对其剂量反应进行了表征。结果表明，固定时间FCM是分析细胞生理和准备应答器的重要工具。 ！23

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