A miniature Couette to generate shear for flow cytometry: studying real-time modulation of intracellular calcium in monocytic cells.
A miniature Couette to generate shear for flow cytometry: studying real-time modulation of intracellular calcium in monocytic cells.
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DOI：
10.1002/cyto.a.21027
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发表时间：
2011-03
期刊：
影响因子：
3.7
通讯作者：
Sklar, Larry A.
中科院分区：
文献类型：
--
作者：
Zwartz, Gordon J.;Chigaev, Alexandre;Foutz, Terry D.;Edwards, Bruce;Sklar, Larry A.
细胞外水动力可能通过改变整合素的构象和亲和力而传递到细胞内部。整合素的激活调节白细胞的募集、细胞的激活和迁移。整合素激活的细胞和分子机制尚不完全清楚，尽管涉及细胞内钙信号。流式细胞术为实时研究细胞内钙信号提供了一种通用的方法。我们报道了一种利用微型Couette产生定义剪切的新方法。测试包括测量悬浮液中剪切诱导的人单核细胞U937细胞内钙信号。Couette外接流式细胞仪，以6PSI(4.1N/m2)加压。细胞在S−1 0~10 0 0μ范围内受到清晰的剪切，并在10 S范围内连续给予1μL/S的FAC扫描。细胞内钙水平和激活的细胞百分比随着剪切时间和强度的增加而增加。
Extracellular hydrodynamic forces may be transmitted to the interior of cells through the alteration of integrin conformation and affinity. Integrin activation regulates leukocyte recruitment, cell activation, and transmigration. The cellular and molecular mechanisms for integrin activation are not precisely known, although intracellular calcium signaling is involved. Flow cytometry offers a versatile way to study intracellular calcium signaling in real-time. We report a novel method to generate defined shear by using a miniature Couette. Testing involved measuring shear induced intracellular calcium signals of human monoblastoid U937 cells in suspension. The Couette was connected externally to a flow cytometer and pressurized at 6 PSI (4.1 N/m2). Cells were subjected to well-defined shear between 0 and 1000 s−1 and delivered continuously within 10 s to a FACScan at 1 μl/s. Intracellular calcium levels and the percentage of cells activated increased as shear increased in duration and intensity.
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