Concentration gradient used in double-stranded DNA separation by capillary electrophoresis

Liang, Dehai; Chu, Benjamin

doi:10.1002/1522-2683(200208)23:16<2602::aid-elps2602>3.0.co;2-x

Cited by 4 publications

(1 citation statement)

References 25 publications

(36 reference statements)

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“…11- 14 Fabrication of gel pore-size gradients in microanalytical systems has been achieved by mimicking approaches commonly used to fabricate macroscale slab gels; 15 that is, by controlling flow rate and filling duration of polymer precursor solutions loaded using dual syringe pumps (i.e., gradient mixers). 16, 17 A variation of the conventional gradient mixer approach used in slab gels was reported by Chen and Chang, 18 wherein electroosmotic flow was used to load polymer precursor solutions into microfluidic channels. Especially relevant to microsystems, both Liang et al 16 and Chen and Chang 18 noted stringent fabrication demands on control of precursor flow rate and filling time, as the total volume of capillary and microdevice separation channels is quite small.…”

Section: Introductionmentioning

confidence: 99%

Photopolymerized diffusion-defined polyacrylamide gradient gels for on-chip protein sizing

Throckmorton

Singh

et al. 2008

Lab Chip

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We report on a facile diffusion-based photopatterning technique for generating linear and non-linear decreasing pore-size gradients in cross-linked polyacrylamide gels. Diffusion of low viscosity polymer precursor solutions and a two-step photopatterning process were used to define the decreasing pore-size gradient gels in a microfluidic format, thus eliminating the need for controlled mixing and delivery of polymer precursor solutions. We present an analytical model of the non-steady state diffusion process and numerically evaluate that model for direct comparison with empirical characterizations of the gradient gels. We show that the analytical model provides an effective means to predict the steepness and linearity of a desired gradient gel prior to fabrication. To assess electrophoretic assay performance in the microfluidic gradient gels, on-chip sizing of protein samples (20-116 kDa) was investigated. Baseline resolution of six proteins was demonstrated in 4 s using 3.5% to 10% polyacrylamide gradient gels. The demonstrated ability to conduct efficient protein sizing in ultra-short separation lengths (0.3 cm) means low applied electric potentials are needed to achieve the electric field strengths required for protein separations. The low required electric potentials relax operating constraints on electrical components, as is especially important for translation of the assay into pre-clinical and clinical settings. The gradient gel fabrication method reported is amenable to adaptation to non-sizing protein assays, as well as integration with upstream sample preparation steps and subsequent orthogonal downstream assays.

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Section: Introductionmentioning

confidence: 99%

Photopolymerized diffusion-defined polyacrylamide gradient gels for on-chip protein sizing

Throckmorton

Singh

et al. 2008

Lab Chip

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Rapid Generation of Biologically Relevant Hydrogels Containing Long‐Range Chemical Gradients

Villa-Uribe

et al. 2009

Adv Funct Materials

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Many biological processes are regulated by gradients of bioactive chemicals. Thus, the generation of materials with embedded chemical gradients may be beneficial for understanding biological phenomena and generating tissue-mimetic constructs. Here we describe a simple and versatile method to rapidly generate materials containing centimeter-long gradients of chemical properties in a microfluidic channel. The formation of chemical gradient was initiated by a passive-pump-induced forward flow and further developed during an evaporation-induced backward flow. The gradient was spatially controlled by the backward flow time and the hydrogel material containing the gradient was synthesized via photopolymerization. Gradients of a cell-adhesion ligand, Arg-Gly-Asp-Ser NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript (RGDS), was incorporated in the poly(ethylene glycol)-diacrylate (PEG-DA) hydrogels to test the response of endothelial cells. The cells attached and spread along the hydrogel material in a manner consistent with the RGDS gradient profile. A hydrogel containing PEG-DA concentration gradient and constant RGDS concentration was also generated. The morphology of cells cultured on such hydrogel changed from round in the lower PEG-DA concentration regions to well-spread in the higher PEG-DA concentration regions. This approach is expected to be a valuable tool to investigate the cell-material interactions in a simple and high-throughput manner and to design graded biomimetic materials for tissue engineering applications.

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Importance of the counterion in optimization of a borate electrolyte system for analyses of anions in samples with complex matrices performed by capillary zone electrophoresis

et al. 2004

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Borate buffers are common background electrolytes for analyses of anions in capillary zone electrophoresis. Usually, sodium borate at a given pH is used and this specification seems to be sufficient for a successful analysis. In this paper, we show that free migration of OH(-) may deteriorate the analysis of a typical anionic analysis of clinical samples due to uncontrolled migration of OH(-) throughout the systems of analyzed zones and may damage the stacking of anionic analytes of interest. We have proven that the use of ammonium borate may remedy the situation where the presence of ammonium may selectively stop the free migration of OH(-) ions, slow down their effective mobility and bring their safe behavior resulting in reproducible stacking of clinically important anions. Results of real analyses of human serum samples confirmed the proposed method and proved that substitution of sodium for ammonium in borate buffers offers reliable analyses of clinical samples having chloride as the bulk component. The experimental results given in this paper are supported also by computer simulation, which can not only support the positive results but also show the dynamics of the separation that is otherwise hidden to any detection possibilities.

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Concentration gradient used in double-stranded DNA separation by capillary electrophoresis

Cited by 4 publications

References 25 publications

Photopolymerized diffusion-defined polyacrylamide gradient gels for on-chip protein sizing

Photopolymerized diffusion-defined polyacrylamide gradient gels for on-chip protein sizing

Rapid Generation of Biologically Relevant Hydrogels Containing Long‐Range Chemical Gradients

Importance of the counterion in optimization of a borate electrolyte system for analyses of anions in samples with complex matrices performed by capillary zone electrophoresis

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