Influence of ignored and well-known zone distortions on the separation performance of proteins in capillary free zone electrophoresis with special reference to analysis in polyacrylamide-coated fused silica capillaries in various buffers

Hydrochloric acid was investigated as a rinsing reagent to remove adsorbed proteins from linear polyacrylamide-coated capillaries for electrophoresis. Three model proteins were used, namely cytochrome c as a basic protein, beta-lactoglobulin as an acidic protein, and beta-casein as a more easily denaturing protein. In order to regenerate capillary surfaces, they have been rinsed for 5 min with 2 M hydrochloric acid, 5 min with water, and then 30 min with buffer after every tenth run. It was found important to perform this regeneration procedure on time. The obtained results show good repeatability of the apparent EOF mobility with percentage RSDs below 3% (n = 60) in various cases. These good results were mainly confirmed in long-term series with more than 200 runs each. Only very high concentrations (175 microM) of beta-lactoglobulin and beta-casein at pH 3.5 gave RSD% values above 5%. For these conditions, the further test of 85% m/m phosphoric acid as rinsing reagent showed a good repeatability of the apparent EOF mobilities as well.

mentioning

confidence: 99%

“…In order to overcome this problem, hydrochloric acid rinsing has been suggested by Mohabbati et al [10]. It was demonstrated that this procedure improved the repeatability of migration times and peak areas in short series.…”

mentioning

confidence: 99%

Reproducible protein analysis by CE using linear polyacrylamide‐coated capillaries and hydrochloric acid rinsing

Suratman

Wätzig

2007

“…Linear polyacrylamide has been used to eliminate EOF in both fused-silica capillary [74][75][76] and channels in PDMS microchips [77]. Stability of the polyacrylamide coating was enhanced by cross-linking [78].…”

Section: Static Wall Coatingmentioning

confidence: 99%

“…However, the composition of BGE affects separation efficiency, too. Several BGEs for CZE of proteins have been compared and 120 mM ammonium acetate and 150 mM ammonium hydroxyacetate, pH 4 were selected as the most promising buffers for CZE of proteins [74,75].…”

Section: Improving Separationmentioning

confidence: 99%

Capillary electrophoresis of proteins 2003–2005

Dolnı́k¹

2006

139

117

This review article with 304 references describes recent developments in CE of proteins, and covers the two years since the previous review (Hutterer, K., Dolník, V., Electrophoresis 2003, 24, 3998-4012) through Spring 2005. It covers topics related to CE of proteins, including modeling of the electrophoretic migration of proteins, sample pretreatment, wall coatings, improving separation, various forms of detection, special electrophoretic techniques such as affinity CE, CIEF, and applications of CE to the analysis of proteins in real-world samples including human body fluids, food and agricultural samples, protein pharmaceuticals, and recombinant protein preparations.

“…CE is a powerful technique for the separation of macromolecules such as proteins and immunocomplexes [14][15][16][17][18][19]. With both superior separation power and high detection sensitivity, CE can separate free antibody or antigen from bound antibody or antigen rapidly, and is especially suitable for immunoassay [20][21][22][23][24][25].…”

Section: Introductionmentioning

confidence: 99%

CE‐based simultaneous liquid‐phase noncompetitive enzyme immunoassay for three tumor markers in human serum using electrochemical detection

Zhang

2007

A method for indirectly detecting horseradish peroxidase (HRP) was described by CE with electrochemical detection. Details of selection for optimum conditions were presented. The detection limit of free HRP was 1.09610 212 M or 0.94 zmol (S/N = 3). A novel CE-based liquid-phase binding noncompetitive enzyme immunoassay (CE-EIA) was developed. In this method, after the noncompetitive immunoreaction in liquid phase, the free enzyme (HRP)-labeled antibody (Ab*) and the bound enzyme-labeled complex (Ag-Ab*) were separated and then the system of HRP catalyzing H 2 O 2 /o-aminophenol (OAP) reaction was adopted. Prostate specific antigen (PSA), carcinoembryonic antigen (CEA), and human chorionic gonadotropin (HCG) in human serum samples were detected without any sample preparation, with the detection limits (S/N = 3) of 0.22, 0.17 and 0.30 ng/mL, respectively. This technique has been successfully applied to detect simultaneously PSA, CEA, and HCG in 12 min, upon adding these three antigens into human serum to simulate patient serum. It proves that the CE-EIA technique proposed could be developed into a sensitive and new method for simultaneous clinical assay of multianalytes.