In‐capillary detection of fast antibody‐peptide binding using fluorescence coupled capillary electrophoresis

Qin, Yuqin; Qiu, Lin; Qin, Haifang; Ding, Shumin; Liu, Li; Teng, Yiwan; Chen, Yao; Wang, Cheli; Jin-chen, LI; Wang, Jianhao; Jiang, Pengju

doi:10.1002/elps.201500429

Cited by 6 publications

(4 citation statements)

References 47 publications

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“…In addition to monitoring of proteins and polypeptide digestion, CE and CEC methods are used also for investigation of aggregation of peptides, formation of peptide covalent adducts and noncovalent complexes . Fast in‐capillary antibody‐peptide binding was monitored by CE coupled with inverted fluorescence microscope equipped with 100 W mercury lamp, an excitation filter (420 ± 20 nm), and a fiberoptic spectrometer attached to the side port . Monoclonal anti‐HA antibody was mixed with HA (influenza hemmaglutinin) tag labeled by 5‐(and‐6)‐carboxyfluorescein inside the capillary.…”

Section: Applicationsmentioning

confidence: 99%

Recent developments in capillary and microchip electroseparations of peptides (2015–mid 2017)

Kašička

2017

Electrophoresis

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The review brings a comprehensive overview of recent developments and applications of high performance capillary and microchip electroseparation methods (zone electrophoresis, isotachophoresis, isoelectric focusing, affinity electrophoresis, electrokinetic chromatography, and electrochromatography) to analysis, microscale isolation, purification, and physicochemical and biochemical characterization of peptides in the years 2015, 2016, and ca. up to the middle of 2017. Advances in the investigation of electromigration properties of peptides and in the methodology of their analysis (sample preseparation, preconcentration and derivatization, adsorption suppression and EOF control, and detection) are described. New developments in particular CE and CEC methods are presented and several types of their applications to peptide analysis are reported: qualitative and quantitative analysis, determination in complex (bio)matrices, monitoring of chemical and enzymatical reactions and physical changes, amino acid, sequence and chiral analysis, and peptide mapping of proteins. Some micropreparative peptide separations are shown and capabilities of CE and CEC methods to provide important physicochemical characteristics of peptides are demonstrated.

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Section: Applicationsmentioning

confidence: 99%

Recent developments in capillary and microchip electroseparations of peptides (2015–mid 2017)

Kašička

2017

Electrophoresis

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“…Feng et al applied QDs in CE immunoassay and separated antibody and complex efficiently [18]. Although CE was used as an efficient tool to study antigen-antibody interaction [12,13,21], the simultaneous detection of assembly/disassembly of multivalent HA tag and anti-HA antibody in a single in-capillary assay poses a significant challenge. Simultaneous detection is supposed to shortening analysis times, reducing the amount of sample required per assay, which sounds useful in practical applications.…”

Section: Introductionmentioning

confidence: 99%

Simultaneous detection of assembly and disassembly of multivalent HA tag and anti‐HA antibody in single in‐capillary assay

Wang

Qin

et al. 2016

Electrophoresis

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Herein, we have developed an in-capillary assay for simultaneous detection of the assembly and disassembly of the multivalent HA tag peptide and antibody. HA tag with hexahistidine at C terminus (YPYDVPDYAG4 H6 , termed YPYDH6 ) was conjugated with quantum dots (QDs) by metal-affinity force to form a multivalent HA tag (QD-YPYDH6 ). QD-YPYDH6 and monoclonal anti-HA antibody (anti-HA) were sequentially injected into the capillary. They were mixed and assembled inside the capillary. The reaction products were online discriminated and detected by fluorescence coupled capillary electrophoresis (CE-FL). For the in-capillary assay, the binding efficiency of the multivalent HA tag and antibody on was influenced by the molar ratio and injection time. Such novel assay could even give out the self-assembly kinetic constant of QDs and YPYDH6 as KD of 34.1 μM with n (binding cooperativeness) of 2.2 by Hill equation. More importantly, the simultaneous detection of the assembly and imidazole (Im) induced disassembly of the QD-YPYDH6 -anti-HA complex was achieved in a single in-capillary assay. Our study demonstrated a new method for the online detection of antigen-antibody interactions.

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“…In‐capillary injection technique can integrate injection, mixing, reaction, separation and detection, greatly improving detection sensitivity . Recently, our group has successfully detected the rapid binding process of anti‐HA antibody and monovalent HA tag using in‐capillary assay by CE–FL , inspiring us to use this method for an in‐depth study of the interaction of multivalent tags with antibodies.…”

Section: Introductionmentioning

confidence: 99%

Investigation of multivalent interactions between conjugate of quantum dots with c‐Myc peptide tag and the anti‐c‐Myc antibody by capillary electrophoresis with fluorescence detection

Wang

Yang

Liu

et al. 2016

J of Separation Science

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Herein, we report an assay for detecting the binding of a multivalent peptide and antibody within a capillary with the use of fluorescence coupled capillary electrophoresis. Quantum dots and a c-Myc tag containing peptide EQKLISEEDLG H were injected sequentially and formed a multivalent quantum dot-EQKLISEEDLG H assembly within the capillary. The efficiency of the quantum dot-peptide self-assembly was affected by the peptide/quantum dot molar ratio, sampling time, and interval time. Finally, the binding of the monoclonal anti-c-Myc antibody and the multivalent quantum dot-EQKLISEEDLG H ligand was studied using an in-capillary assay. The microscopic dissociation constant for the self-assembly of monoclonal anti-c-Myc antibody and quantum dot-EQKLISEEDLG H was determined to be 14.1 μM with a stoichiometry of the peptide-antibody complex of 1.7 determined after fitting this to the Hill equation. This method can be further extended to detect a wide range of biomolecule-biomolecule binding interactions.

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In‐capillary detection of fast antibody‐peptide binding using fluorescence coupled capillary electrophoresis

Cited by 6 publications

References 47 publications

Recent developments in capillary and microchip electroseparations of peptides (2015–mid 2017)

Recent developments in capillary and microchip electroseparations of peptides (2015–mid 2017)

Simultaneous detection of assembly and disassembly of multivalent HA tag and anti‐HA antibody in single in‐capillary assay

Investigation of multivalent interactions between conjugate of quantum dots with c‐Myc peptide tag and the anti‐c‐Myc antibody by capillary electrophoresis with fluorescence detection

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