Over 1000-fold protein preconcentration for microliter-volume samples at a pH junction using capillary electrophoresis

Nesbitt, Chandra A.; Lo, Josephine T.-M.; Yeung, Ken K.‐C.

doi:10.1016/j.chroma.2004.09.081

Cited by 45 publications

(54 citation statements)

References 21 publications

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“…Nesbitt et al [154] presented a method for the preconcentration of proteins using a pH boundary created through the neutralisation of acetate (low pH buffer) with ammonium (high pH buffer). This is distinctly different from previous approaches in that the pH boundary is created through neutralisation of a weak base with a weak acid, while in all previous reports the boundary has been created using only buffers containing weak acids.…”

Section: Dynamic Ph Junctionmentioning

confidence: 99%

Recent advances in enhancing the sensitivity of electrophoresis and electrochromatography in capillaries and microchips

2007

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Poor sensitivity is considered to be one of the major limitations of electrophoretic separation methods, particularly when compared to traditional liquid chromatographic techniques. To address this issue, various in-line preconcentration techniques have been developed over the past 15 years, ranging in power and complexity, and there are now a number of well understood approaches routinely capable of providing a 10,000- to 100,000-fold increase in sensitivity, as well as several that can be pushed above a million. Furthermore, these have been achieved with particularly troublesome and often difficult samples, such as those having high salinity from a biological or environmental origin. This review will discuss the most common methods for improving the sensitivity of CE, CEC and microchip version of these, with particular attention to those approaches developed over the last five years.

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Section: Dynamic Ph Junctionmentioning

confidence: 99%

Recent advances in enhancing the sensitivity of electrophoresis and electrochromatography in capillaries and microchips

2007

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“…The first uses interaction between the analytes and a pseudostationary phase to induce the preconcentration, and the second is based on pH differences along the separation path [12][13][14][15][16][17][18][19]. Although a high sensitivity enhancement can be reached when one of the above-mentioned methods is integrated in CZE, even higher SEFs can be achieved when combining these techniques.…”

Section: Introductionmentioning

confidence: 99%

Electrokinetic supercharging for highly efficient peptide preconcentration in capillary zone electrophoresis

2008

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Electrokinetic supercharging has been integrated in CZE for the development of a highly sensitive methodology for protein tryptic digest analysis. A careful choice of the experimental conditions led to sensitivity enhancement factors between 1000 and 10 000 whilst maintaining a satisfactory resolution. Peptides in the low nanomolar concentration range have been detected despite the use of the poorly sensitive UV absorbance detection mode. The buffer system used in this study is fully suitable for coupling CE to MS.

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“…Numerous online concentration technologies, such as field amplification [26], ITP [27], sweeping [28], and pH junction [29], have been reported for the enhancement of detection sensitivity in CE. In these methods, discontinuous buffer systems between sample and BGE zones are used to build the difference in electric field strength and analyte mobility.…”

Section: Introductionmentioning

confidence: 99%

Online concentration and separation of basic proteins using a cationic polyelectrolyte in the presence of reversed electroosmotic flow

Tseng

2006

Electrophoresis

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We report an online concentration and separation method for basic proteins using poly(diallyldimethylammonium chloride) (PDDA) solutions in the presence of reversed EOF. Using a capillary dynamically coated with 2% PDDA containing 0.1 M NaCl and filled with 1.2% PDDA under neutral conditions (10 mM phosphate, pH 7.0), we have demonstrated the separation of six basic proteins with peak efficiencies ranging from 175 000 to 616 000 plates/m and RSDs of migration time less than 0.4%. Additionally, high-speed separation of six basic proteins (<7 min) was achieved using a short capillary filled with 0.6% PDDA solutions. Under injection of the large-volume sample (210 nL), the LODs at S/N of 3 for basic proteins are down to nanomolar range. For example, the LOD for lysozyme is 1.2 nM, which is a 260-fold sensitivity enhancement compared with conventional injection method. The proposed method has been applied to the stacking of lysozyme in human saliva samples. Without any pretreatment, we also demonstrated the capability of this method to detect low amounts of peptide samples through the stacking of tryptic peptide of myoglobin. The experimental results indicate that our proposed method has great potential for use in clinical diagnosis and proteomics applications.

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Over 1000-fold protein preconcentration for microliter-volume samples at a pH junction using capillary electrophoresis

Cited by 45 publications

References 21 publications

Recent advances in enhancing the sensitivity of electrophoresis and electrochromatography in capillaries and microchips

Recent advances in enhancing the sensitivity of electrophoresis and electrochromatography in capillaries and microchips

Electrokinetic supercharging for highly efficient peptide preconcentration in capillary zone electrophoresis

Online concentration and separation of basic proteins using a cationic polyelectrolyte in the presence of reversed electroosmotic flow

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