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A rapid microfluidic technique for integrated viability determination of adherent single cells

Shijun Xu (Institutionen för kemi och kemiteknik, Fysikalisk kemi) ; Anna Kim (Institutionen för kemi och kemiteknik, Fysikalisk kemi) ; Gavin Jeffries (Institutionen för kemi och kemiteknik, Fysikalisk kemi) ; Aldo Jesorka (Institutionen för kemi och kemiteknik, Fysikalisk kemi)
Analytical and Bioanalytical Chemistry (1618-2642). Vol. 407 (2015), 5, p. 1295-1301.
[Artikel, refereegranskad vetenskaplig]

Here, we report on a novel protocol for determining the viability of individual cells in an adherent cell culture, without adversely affecting the remaining cells in the sample. This is facilitated using a freestanding microfluidic perfusion device, the Multifunctional Pipette (MFP), which generates a virtual flow cell around selected single cells. We investigated the utility on four different cell lines, NG108-15, HEK 293, PC12, and CHO, and combined the assay with a cell poration experiment, in which we apply the pore-forming agent digitonin, followed by fluorescein diphosphate, a pre-fluorescent substrate for alkaline phosphatase, in order to monitor intracellular enzyme activity. The cell viability was instantly assessed through simultaneous perfusion with fluorescein diacetate (FDA) and propidium iodide (PI), both being dispensed through the same superfusion device used to porate and deliver the enzyme substrate. In this fluorescence assay, viable and non-viable cells were distinguished by their green and red emission, respectively, within 10 s. In addition, the enzyme activity was monitored over time as a secondary test for cellular activity. Our findings demonstrate that this microfluidic technology-assisted approach is a facile, rapid, and reliable means to determine the viability in single-cell experiments and that viability studies can be performed routinely alongside typical substrate delivery protocols. This approach would remove the need for global cell viability testing and would enable viability studies of only the cells under experimental analysis.

Nyckelord: Adherent single cells, Viability, Multifunctional Pipette, FDA and PI



Denna post skapades 2015-03-11. Senast ändrad 2016-07-12.
CPL Pubid: 213663

 

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Institutioner (Chalmers)

Institutionen för kemi och kemiteknik, Fysikalisk kemi

Ämnesområden

Biokemi och molekylärbiologi

Chalmers infrastruktur