THE DISTRIBUTION OF INTRACELLULAR CALCIUM CHELATOR (FURA-2) IN A POPULATION OF INTACT HUMAN RED-CELLS
THE DISTRIBUTION OF INTRACELLULAR CALCIUM CHELATOR (FURA-2) IN A POPULATION OF INTACT HUMAN RED-CELLS
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DOI：
10.1016/0005-2736(93)90172-v
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发表时间：
1993-05-14
期刊：
影响因子：
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通讯作者：
EISINGER, J
中科院分区：
文献类型：
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作者：
LEW, VL;ETZION, Z;EISINGER, J
用荧光定量显微镜观察Fura-2的1-2.5mmo1-2.5mmo1/L细胞负载的红细胞，观察掺入的游离螯合剂在单个细胞内和细胞内的分布。胞质血红蛋白使Fura-2的有效荧光产额猝灭约100倍。所有的红细胞在380 nm激发时都有荧光，它们在510 nm处发射的荧光强度约为平均强度的+/-20%，表明所掺入的螯合剂在细胞中的分布相当均匀。负载这些高水平Fura-2的红细胞保持了它们的双凹形状，在415 nm处的透射图与它们的Fura-2荧光图的比较表明，Fura-2在整个细胞质中的浓度也是均匀的。这些结果验证了使用非荧光掺入的钙离子螯合剂的早期实验中所做的假设，并证明了Fura-2和完整红细胞的钙成像的可行性，尽管探针荧光被血红蛋白相当程度地猝灭。
Using quantitative fluorescence microscopy of red cells loaded non-disruptively with 1-2.5 mmol/l cells of fura-2, we examined the distribution of the incorporated free chelator among and within individual cells. Cytoplasmic hemoglobin quenched the effective fluorescence yield of fura-2 by a factor of about 100. All red cells were found to fluoresce upon excitation at 380 nm, and the fluorescence intensities they emitted at 510 nm were approximately +/- 20% about the mean intensity, indicating a fairly uniform distribution of incorporated chelator among the cells. Red cells loaded with these high levels of fura-2 retained their biconcave shape, and a comparison between their transmission images at 415 nm and their fura-2 fluorescence images suggests that the concentration of fura-2 was also uniform throughout the cytosol. These results validate assumptions made in earlier experiments with non-fluorescent incorporated Ca2+ chelators, and demonstrate the feasibility of fura-2 and Ca2+ imaging of intact red cells, despite considerable quenching of probe fluorescence by hemoglobin.