Automated analysis of individual particles using a commercial capillary electrophoresis system

Ahmadzadeh, Hossein; Dua, Rajat A.; Presley, Andrew D.; Arriaga, Edgar A.

doi:10.1016/j.chroma.2004.12.018

Cited by 29 publications

(23 citation statements)

References 27 publications

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“…Using these methods, cell viability can be determined by analyzing populations of cells, but not based on the state of individual cells. In recent years, Arriaga and his colleagues used CE for separation and detection of individual particles including nuclei [5], mitochondria [6-9, 23, 24], acidic organelles [10][11][12][13][14], liposomes [25], microspheres [26], and latex beads [27]. In all these reported experiments, CZE mode was used to analyze individual particles.…”

Section: Introductionmentioning

confidence: 99%

Continuous intact cell detection and viability determination by CE with dual‐wavelength detection

Ren

Zhang

et al. 2010

Electrophoresis

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We introduce here a method for continuous intact cell detection and viability determination of individual trypan blue stained cells by CE with ultraviolet-visible dual-wavelength detection. To avoid cell aggregation or damage during electrophoresis, cells after staining were fixed with 4% formaldehyde and were continuously introduced into the capillary by EOF. The absorbance of a cell at 590 nm was used to determine its viability. An absorbance of two milli-absorbance unit at 590 nm was the clear cut-off point for living and dead Hela cells in our experiments. Good viability correlation between the conventional trypan blue staining assay and our established CE method (correlation coefficient, R(2)=0.9623) was demonstrated by analysis of cell mixtures with varying proportions of living and dead cells. The CE method was also used to analyze the cytotoxicity of methylmercury, and the results were in good agreement with the trypan blue staining assay and 3-(4,5-dimethyl-2-thiazyl)-2,5-diphenyl-2H-tetrazolium bromide methods. Compared with the 3-(4,5-dimethyl-2-thiazyl)-2,5-diphenyl-2H-tetrazolium bromide method, our established CE method can be easily automated to report cell viability based on the state of individual cells. Tedious manual cell counting and human error due to investigator bias can be avoided by using this method.

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Section: Introductionmentioning

confidence: 99%

Continuous intact cell detection and viability determination by CE with dual‐wavelength detection

Ren

Zhang

et al. 2010

Electrophoresis

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“…This theory was later adopted for CE by Ahmadzadeh et al [37] and utilized to evaluate the peak overlap in an electropherogram of latex microparticles. The basis of SOT centers around the following equation [35]:…”

Section: Oversamplingmentioning

confidence: 99%

“…Given the definition of e 2a the probability that a peak from one event will not overlap with either the event before it or the event after it is given by [37] p SCP = (e 2a )(e 2a ) = e 22a (7) where p SCP is the probability that any one peak is the result of a single component, or event. The number of single component peaks is then given by…”

Section: Oversamplingmentioning

confidence: 99%

“…Using the SOT procedure developed previously [37], the probability that any individual peak will actually be composed of multiple events in the 2 s window is 28% (three overlapping peaks) and 24% (two overlapping peaks) when the number of events equals the N OT for rat liver and 143B mitochondria, respectively. Considering that this is only a 2 s portion of the electropherogram, this shows that the number of multiple component peaks is a major concern in analysis of individual mitochondrial events in microfluidic devices.…”

Section: Oversamplingmentioning

confidence: 99%

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Determining under‐ and oversampling of individual particle distributions in microfluidic electrophoresis with orthogonal laser‐induced fluorescence detection

et al. 2008

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This report investigates the effects of sample size on the separation and analysis of individual biological particles using microfluidic devices equipped with an orthogonal LIF detector. A detection limit of 17 6 1 molecules of fluorophore is obtained using this orthogonal LIF detector under a constant flow of fluorescein, which is a significant improvement over epifluorescence, the most common LIF detection scheme used with microfluidic devices. Mitochondria from rat liver tissue and cultured 143B osteosarcoma cells are used as model biological particles. Quantile-quantile (q-q) plots were used to investigate changes in the distributions. When the number of detected mitochondrial events became too large (.72 for rat liver and .98 for 143B mitochondria), oversampling occurs. Statistical overlap theory is used to suggest that the cause of oversampling is that separation power of the microfluidic device presented is not enough to adequately separate large numbers of individual mitochondrial events. Fortunately, q-q plots make it possible to identify and exclude these distributions from data analysis. Additionally, when the number of detected events became too small (,55 for rat liver and ,81 for 143B mitochondria) there were not enough events to obtain a statistically relevant mobility distribution, but these distributions can be combined to obtain a statistically relevant electrophoretic mobility distribution.

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“…This is primarily due to the much lower prices and smaller size of HV modules below 10 kV. Typical voltage accuracies of 0.1% of maximum voltage, resolutions in the µV range and stabilities of less than 50 mV peak to peak can be found from commercial suppliers [153][154][155]. When integrating these into a P-CE system, typically the power supply can be programmed to pro-vide the required voltage profile during detection.…”

Section: 7mentioning

confidence: 99%

Portable capillary-based (non-chip) capillary electrophoresis

Ryvolová

Preisler

Brabazon

et al. 2010

TrAC Trends in Analytical Chemistry

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Miniaturized, portable instrumentation has been gaining popularity in all areas of analytical chemistry. Capillary electrophoresis (CE), due to its main strengths of high separation efficiency, relatively short analysis time and low consumption of chemicals, is a particularly suitable technique for use in portable analytical instrumentation. In line with the general trend in miniaturization in chemistry utilizing microfluidic chips, the main thrust of portable CE (P-CE) systems development is towards chip-based miniaturized CE. Despite this, capillary-based (non-chip) P-CE systems have certain unmatched advantages, especially in the relative simplicity of the regular cylindrical geometry of the CE capillary, maximal volume-to-surface ratio, no need to design and to fabricate a chip, the low costs of capillary compared to chip, and better performance with some detection techniques. This review presents an overview of the state of the art of P-CE and literature relevant to future developments. We pay particular attention to the development and the potential of miniaturization of functional parts for P-CE. These include components related to sample introduction, separation and detection, which are the key elements in P-CE design. The future of P-CE may be in relatively simple, rugged designs (e.g., using a short piece of capillary fixed to a chip-sized platform on which injection and detection parts can be mounted). Electrochemical detection is well suited for miniaturization, so is probably the most suitable detection technique for P-CE, but optical detection is gaining interest, especially due to miniaturized light sources (e.g., light-emitting diodes).

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Automated analysis of individual particles using a commercial capillary electrophoresis system

Cited by 29 publications

References 27 publications

Continuous intact cell detection and viability determination by CE with dual‐wavelength detection

Continuous intact cell detection and viability determination by CE with dual‐wavelength detection

Determining under‐ and oversampling of individual particle distributions in microfluidic electrophoresis with orthogonal laser‐induced fluorescence detection

Portable capillary-based (non-chip) capillary electrophoresis

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