Capillary electrophoresis of single cells: observation of two compartments of neurotransmitter vesicles

Kristensen, Helle Kjærgaard; Lau, Yau Yi; Ewing, Andrew G.

doi:10.1016/0165-0270(94)90009-4

Cited by 80 publications

(45 citation statements)

References 16 publications

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“…Krilov et al showed that single cell analysis with CE-LIFD gives more accurate metabolic information than cellular extract analysis [152]. The CA have been analyzed by CE-EC or CE-LIFD in different single cells [133][134][135][139][140][141]153]. Steroids were found in single R2C cell while progesterone secretion was monitored from culture cells and subsequently in single cells.…”

Section: Ce and Single Cell Analysismentioning

confidence: 99%

Biomedical applications of capillary electrophoresis with laser‐induced fluorescence detection

Páez

Hernández

2001

Biopharm & Drug Disp

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Capillary electrophoresis (CE) is a high-efficiency analytical technique that has had a great impact as a tool in biomedical research, clinical and forensic practice in the last ten years. Only in one of the applications, the DNA analysis, it has had an explosive exponential growth in the last few years. This impact is expressed in an enormous amount of CE articles and many reviews. The CE advantages with respect to other analytical techniques: the required very small sample volume, rapid analysis, great resolution power and low costs, have made this technique ideal for the analysis of a numerous endogenous and exogenous substances present in biological fluids. The different modes of CE have been coupled to different detection techniques such as UV-absorbance, electrochemical, mass spectrometry and laser-induced fluorescence detection (LIFD) to detect different nature and molecular size separated analytes. This review focuses mostly on the applications of CE-LIFD, to measure drugs and endogenous neuroactive substances such as amino acids and monoamines, especially in microdialysis samples from experimental animals and humans. CE-LIFD trends are discussed: automated faster analysis with capillary array systems, resolution power improvement, higher detection sensitivity, and CE systems miniaturization for extremely small sample volume, in order to make CE easier and affordable to the lab bench or the clinical bed.

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Section: Ce and Single Cell Analysismentioning

confidence: 99%

Biomedical applications of capillary electrophoresis with laser‐induced fluorescence detection

Páez

Hernández

2001

Biopharm & Drug Disp

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“…Hogan and Yeung [12] were the first to inject intact single cells into a capillary for component characterization. Ewing [13][14][15] has used subcellular sampling to characterize components in localized regions of cells, and thus has simplified the tremendous complexity exhibited in these biological systems. Arriaga [16,17] has also achieved subcellular sampling of mitochondria organelles using LIF detection.…”

Section: Introductionmentioning

confidence: 99%

Cell cycle‐dependent characterization of single MCF‐7 breast cancer cells by 2‐D CE

et al. 2007

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The composition of single MCF-7 breast cancer cells is characterized using 2-D CE. Individual MCF-7 cells were aspirated into a 30 mum inner diameter fused-silica capillary and lysed by contact with an SDS-containing buffer. Proteins and other primary amines were fluorescently labeled on-column using the fluorogenic dye 3-(2-furoyl)quinoline-2-carboxaldehyde. Labeled components were separated first according to molecular weight using capillary sieving electrophoresis (CSE) and then by MEKC. Analytes were detected in a sheath-flow cuvette using LIF. The expression profiles for MCF-7 cellular homogenate and a single MCF-7 cell are compared. As a proof-of-principle investigation, variation in expression was also compared within and between G1 and G2/M cell cycle phases for MCF-7 cells. Following their treatment with the viable nuclear stain Hoechst 33342, MCF-7 cells were sorted by flow cytometry on the basis of their ploidy. Sorted cells were then analyzed by 2-D CE. The degree of variability was >2.5 times larger between cells of different phases than between cells of the same phase. In typical 1 h 2-D CE separations using MCF-7 cells, over 100 components are resolved.

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“…This can be achieved by using surfaces, to which cells do not adhere, or by using suspended cells such as hemopoietic stem cells. Four techniques of cell lysis inside the capillary have been described: (i) ultrasonic treatment [20], (ii) high-electric field [16,19], (iii) lowionic-strength solution [21], and (iv) a surfactant [8,17,22]. Using a surfactant is a simple and efficient way.…”

Section: Introductionmentioning

confidence: 99%

“…Using a surfactant is a simple and efficient way. Cell lysis was demonstrated using a surfactant in the run buffer [8,17] and by a plug of a surfactant injected after the cell [22]. If a surfactant is used to lyse cells in chemical cytometry of protein-DNA interactions, it must not affect the stability of protein-DNA complexes.…”

Section: Introductionmentioning

confidence: 99%

Selection of surfactants for cell lysis in chemical cytometry to study protein‐DNA interactions

et al. 2006

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Protein-DNA interactions play a defining role in many cellular processes. Studying such interactions at the single-cell level is important and challenging. Here we make the first step toward achieving this goal with chemical cytometry. Chemical cytometry utilizes capillary separation for detailed chemical analyses of single cells. The cell is injected into a capillary, lysed, and its components are analyzed by CE or capillary chromatography with highly sensitive detection. In order to apply chemical cytometry to studies of protein-DNA interactions, cell lysis must not destroy protein-DNA complexes. Surfactants represent the most practical means of cell lysis inside the capillary. This work aimed at finding surfactants and lysis conditions that do not destroy protein-DNA complexes. We studied three groups of surfactants--ionic, zwitterionic, and nonionic--with respect to their ability to lyse the cell membrane without significantly influencing the stability of protein-DNA complexes. Nonequilibrium CE of equilibrium mixtures with surfactants in the equilibrium mixtures and in the run buffer was used to measure the equilibrium constant, K(d), and rate constant, k(off), of protein-DNA complex dissociation. We found that nonionic surfactants worked best: they lyse the plasma membrane without significantly influencing K(d), k(off), or the EOF. This work creates the foundation for studies of protein-DNA interactions in single cells by chemical cytometry.

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Capillary electrophoresis of single cells: observation of two compartments of neurotransmitter vesicles

Cited by 80 publications

References 16 publications

Biomedical applications of capillary electrophoresis with laser‐induced fluorescence detection

Biomedical applications of capillary electrophoresis with laser‐induced fluorescence detection

Cell cycle‐dependent characterization of single MCF‐7 breast cancer cells by 2‐D CE

Selection of surfactants for cell lysis in chemical cytometry to study protein‐DNA interactions

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