Concomitant detection of CYP1A1 enzymatic activity and CYP1A1 protein in individual cells of a human urothelial cell line using a bilayer microfluidic device

Abstract

To understand molecular networking at the cellular level, analyses of processes and effects at the single-cell level are most appropriate. Usual biochemical or molecular biological analyses are based on integrated signals of numerous cells which differ, however, in their expression and activity profiles. Here we show that it is possible to determine different types of properties of individual cells by means of a specifically designed microfluidic device. As part of investigations to characterize the human urothelial cell line 5637 as a potential model system for studies of toxic and carcinogenic effects on urothelial cells, we use this cell line to assign cytochrome P450 activity, and expression of the enzymes involved, to individual cells. It is shown that the cell population is very heterogeneous with respect to the extent and kinetics of CYP1A1-dependent ethoxyresorufin O-deethylase (EROD). This is also true for the cells' CYP1A1 protein content. With some exceptions, the EROD activity largely coincides with the presence of CYP1A1 protein in the cells. The results obtained with the microfluidic device are promising and open up new perspectives with regard to multi-property determinations in individual cells and to studies focusing on the biochemical and molecular heterogeneity of cells.

A simple and fast microfluidic approach of same-single-cell analysis (SASCA) for the study of multidrug resistance modulation in cancer cells

“…14,16,17 Additionally, physicochemical modelling of biological processes also demands single-cell data. 18 These single-cell applications include intracellular signalling, [19][20][21] pathogen identification, 22 myocyte contraction, 19,[23][24][25] drug discovery, 23,26 patch-clamp recording, 27 multidrug resistance, 28 cell nanosurgery, 29 electroporation, 30 genetic analysis, 31,22 protein analysis, 32 and so on.…”

Cancer Stem Cells - The Cutting Edge

“…Additionally, physicochemical modelling of biological processes also demands data to be obtained from a single cell (El-Ali et al, 2006). To date, the single-cell applications using the microfluidic devices include intracellular signalling (Wheeler et al, 2003) (Li et al, 2004), pathogen identification (Zeng et al, 2010), myocyte contraction ) ) (Klauke et al, 2003), drug discovery , patch-clamp recording (Chen and Folch, 2006), multidrug resistance (Li et al, 2008), cell nanosurgery (Jeffries et al, 2007), electroporation (Ryttsen et al, 2000), genetic analysis (Liu et al, 2010) (Hong et al, 2004), protein analysis (Schumann et al, 2008), forensic analysis (Liu et al, 2010), cell culture , photobleaching of cellular fluorescence (Peng and Li, 2005), and so on. Recently, we have developed a microfluidic single-cell analysis approach for the study of multidrug resistance modulation by real-time monitoring of drug efflux in MDR cancer cells (Li et al, 2008).…”

Monitoring induced gene expression of single cells in a multilayer microchip

“…Additionally, the delivery of stimulants can be spatially resolved using multilaminar flows allowing the formation of chemical gradients. Hence, the two-layer microfluidic platform allows a On the basis of previous designs [35][36][37], we improved the cell handling by implementation of a cell culture chamber with a trap array in the top channel. Precise cell positioning simplifies the currently semiautomated analysis of the individual mammalian cells, and will pave the way for fully automated data analysis in the future.…”