Immunoaffinity CE for Proteomics Studies.

Guzman, Norberto A.; Phillips, Terry M.

doi:10.1021/ac053325c

Cited by 82 publications

(88 citation statements)

References 32 publications

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“…After an equilibration time of 10 min to ensure antigen binding, we measured the concentration of the remaining NT-proBNP. Secondly, the purified FabЈ 1-21 fragments were coupled to beads following the approach of Guzman et al (20,21 ), with the exception that amino-derivatized silica beads (bead size, 7 m; pore size, 30 nm) were used. After washing the silica beads (16 g/L) 3 times with sodium phosphate buffer (50 mmol/L, pH 7.2), we covalently bound 5 g/L sulfo-succinimidyl 4-(N-maleimidomethyl)cyclohexane-1-carboxylate (S-SMCC)(Sigma-Aldrich) to the free amino groups for 1 h at 30°C.…”

Section: Affinity Chromatographymentioning

confidence: 99%

Analysis of Circulating Forms of proBNP and NT-proBNP in Patients with Severe Heart Failure

et al. 2008

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Section: Affinity Chromatographymentioning

confidence: 99%

Analysis of Circulating Forms of proBNP and NT-proBNP in Patients with Severe Heart Failure

et al. 2008

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“…In the field of electrokinetically driven separation, online immunoaffinity CE (IA-CE) has opened the way to more straightforward protocols [1][2][3][4][5]. It involves the online hybridization of two technologies: immunoaffinity extraction and CE.…”

Section: Introductionmentioning

confidence: 99%

Capillary electrophoresis immunoassay using magnetic beads

Chen¹,

Busnel²,

Gassner³

et al. 2008

Electrophoresis

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Protein A-coated magnetic beads (0.3 mm) have been trapped in a small portion of a neutrally coated capillary (50 mm id). Anti-b-lactoglobulin (b-LG) antibodies have then been immobilized on the beads through strong affinity with protein A to subsequently capture b-LG from model or real samples. Once the immunocomplexes formed at physiological pH, a discontinuous buffer system has been used to release the partners and preconcentrate them by transient ITP. The antigens and antibodies have finally been separated by CZE and detected by UV absorbance. An LOQ of 55 nM has been achieved. This methodology has been applied to quantify native b-LG in pasteurized and ultra-high-temperature-treated bovine milk. All the described procedures, including immunosorbent preparation, sample extraction, cleanup, preconcentration, and separation are completely automated on a commercial CE instrument. As this CE immunoassay method is simple, rapid, selective, and sensitive, it should be a practical and attractive technology for the analysis of complicated biological samples.

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“…CE can be enhanced further by coupling its separation power to the selectivity of affinity or immunoaffinity ligands such as lectins or antibodies. These agents can act not only as selective capture molecules but also as preanalytical concentrators [34][35][36]. Immobilized antibodies, either in preanalytical; concentrators or directly immobilized into the capillary itself have been applied to the analysis of a number of analytes including molecules of clinical relevance or interest [37][38][39][40][41].…”

Section: Discussionmentioning

confidence: 99%

Analysis of inflammatory biomarkers from tissue biopsies by chip‐based immunoaffinity CE

Phillips

Wellner

2007

Electrophoresis

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To aid in the biochemical analysis of human skin biopsies, a semiautomatic chip-based CE system has been developed for measuring inflammatory biomarkers in microdissected areas of the biopsy. Following solubilization of the dissected tissue, the desired biomarkers were isolated by immunoaffinity capture using a panel of 12 antibodies, immobilized on a disposable glass fiber disk, within the extraction port of the chip. The captured analytes were labeled with a 635 nm light-emitting laser dye and electroeluted into the separation channel. Electrophoretic separation of all of the analytes was achieved in 2.2 min with quantification of each peak being performed by online LIF detection and integration of each peak area. Comparison of the results obtained from the chip-based system to those obtained using commercially available high-sensitivity immunoassays demonstrated that the chipbased assay provides a fast, accurate procedure for studying the concentrations of inflammatory biomarkers in complex biological materials. The degree of accuracy and precision achieved by the chip-based CE is comparable to conventional immunoassays and the system is capable of analyzing circa six samples per hour. With the ever-expanding array of antibodies that are commercially available, this chip-based system can be applied to a wide variety of different biomedical analyses.

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Immunoaffinity CE for Proteomics Studies.

Cited by 82 publications

References 32 publications

Analysis of Circulating Forms of proBNP and NT-proBNP in Patients with Severe Heart Failure

Analysis of Circulating Forms of proBNP and NT-proBNP in Patients with Severe Heart Failure

Capillary electrophoresis immunoassay using magnetic beads

Analysis of inflammatory biomarkers from tissue biopsies by chip‐based immunoaffinity CE

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