Determination of neurotransmitters in PC 12 cells by microchip electrophoresis with fluorescence detection

Shi, Baoxian; Huang, Wei‐Hua; Cheng, Jie

doi:10.1002/elps.200600615

Cited by 25 publications

(17 citation statements)

References 28 publications

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“…An average human erythrocyte contains 60 ± 30 amol of glutathione; 12 even a single molecule in a 60 fL erythrocyte will be present at about a nanomolar concentration, so that lower concentration measurements are not needed. Similarly, a PC12 cell includes 3.5 ± 3 fmol of glutamic acid, 13 and larger neurons can contain femtomole levels of select neuropeptides. 14 …”

Section: What Are the Metabolome And Peptidome?mentioning

confidence: 99%

Profiling metabolites and peptides in single cells

et al. 2011

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Metabolites and peptides play important roles in almost every aspect of cell function. Their intracellular levels and spatial localizations reflect the state of each cell and its relationship to its surrounding environment. Moreover, their levels and dynamics are indicative of normal or pathological cellular conditions. Bioanalytical technologies for microanalysis are able to qualitatively and quantitatively characterize subsets of peptides and metabolites from individual microorganism, plant and animal cells. Highlighted here are the established and evolving strategies for characterization of the metabolome and peptidome of single cells. Focused studies of the chemical composition of individual cells and their networks promise to provide a greater understanding of cellular fate, function, and homeostatic balance. Single cell bioanalytical microanalysis has also become increasingly valuable for examining cellular heterogeneity, particularly in the fields of neuroscience, stem cell biology, and developmental biology.

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Section: What Are the Metabolome And Peptidome?mentioning

confidence: 99%

Profiling metabolites and peptides in single cells

et al. 2011

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“…DA and Glu are two major neurotransmitters in PC-12 neuronal cells [40]. DA is involved in initiating many behavioral responses towards various stimuli, and Glu is the major excitatory neurotransmitter in the mammalian central nervous system [41, 42].…”

Section: Resultsmentioning

confidence: 99%

A microchip electrophoresis-mass spectrometric platform with double cell lysis nano-electrodes for automated single cell analysis

Zhao

et al. 2016

Journal of Chromatography A

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Capillary electrophoresis-based single cell analysis has become an essential approach in researches at the cellular level. However, automation of single cell analysis has been a challenge due to the difficulty to control the number of cells injected and the irreproducibility associated with cell aggregation. Herein we report the development of a new microfluidic platform deploying the double nano-electrode cell lysis technique for automated analysis of single cells with mass spectrometric detection. The proposed microfluidic chip features integration of a cell-sized high voltage zone for quick single cell lysis, a microfluidic channel for electrophoretic separation, and a nanoelectrospray emitter for ionization in MS detection. Built upon this platform, a microchip electrophoresis - mass spectrometric method (MCE-MS) has been developed for automated single cell analysis. In the method, cell introduction, cell lysis, and MCE-MS separation are computer controlled and integrated as a cycle into consecutive assays. Analysis of large numbers of individual PC-12 neuronal cells (both intact and exposed to 25 mM KCl) was carried out to determine intracellular levels of dopamine (DA) and glutamic acid (Glu). It was found that DA content in PC-12 cells was higher than Glu content, and both varied from cell to cell. The ratio of intracellular DA to Glu was 4.20 ± 0.8 (n=150). Interestingly, the ratio drastically decreased to 0.38 ± 0.20 (n= 150) after the cells are exposed to 25 mM KCl for 8 min, suggesting the cells released DA promptly and heavily while they released Glu at a much slower pace in response to KCl-induced depolarization. These results indicate that the proposed MCE-MS analytical platform may have a great potential in researches at the cellular level.

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“…Good linearity has been obtained by plotting peak area versus analyte concentration, with correlation coefficients larger than 0.9957. The LODs of the labeled amines were in the range from 0.65 nM for Gly to 73 nM for dopamine (DA) (S/N 5 3), which are equivalent or better than the detection limits obtained via other methods for AAs and catecholamines [21,22,33].…”

Section: Validation Of the Methodsmentioning

confidence: 98%

Separation of free amino acids and catecholamines in human plasma and rabbit vitreous samples using a new fluorogenic reagent 3‐(4‐bromobenzoyl)‐2‐quinolinecarboxaldehyde with CE‐LIF detection

Zhang

Wang

2009

Electrophoresis

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A sensitive and efficient analysis of amino acids and catecholamines is currently presented with 3-(4-bromobenzoyl)-2-quinolinecarboxaldehyde as a fluorogenic derivatization reagent using CE separation with LIF detection. For good derivatization conditions, the reagent concentrations, pH value, temperature, and reaction time were explored, which were followed by the derivatization reaction in stable yield. The optimal running buffer was composed of mixtures involving 120 mM, pH 9.1 boric acid, 38.5 mM SDS, and 19% ACN v/v. The LOD (S/N=3) was found as low as 0.65 nM. The proposed method was validated by the two-order-magnitude linearity and correlation coefficient ranging from 0.9957 to 0.9998. The accuracy and specificity of this assay were also assured from the spiking of real samples with a standard known concentration. In order to demonstrate its wide range of applications, the method has been applied to the analysis of both human plasma and rabbit vitreous samples.

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Determination of neurotransmitters in PC 12 cells by microchip electrophoresis with fluorescence detection

Cited by 25 publications

References 28 publications

Profiling metabolites and peptides in single cells

Profiling metabolites and peptides in single cells

A microchip electrophoresis-mass spectrometric platform with double cell lysis nano-electrodes for automated single cell analysis

Separation of free amino acids and catecholamines in human plasma and rabbit vitreous samples using a new fluorogenic reagent 3‐(4‐bromobenzoyl)‐2‐quinolinecarboxaldehyde with CE‐LIF detection

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