Affinity capillary electrophoresis with laser induced fluorescence detection for thrombin analysis using nuclease-resistant RNA aptamers

Líu, Hao; Bai, Yunlong; Wang, Hailin; Zhao, Qiang

doi:10.1016/j.chroma.2016.11.011

Cited by 14 publications

(10 citation statements)

References 33 publications

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“…Fluorescein isothiocyanate (FITC)-labeled thrombin aptamer was obtained from Aptamer Sciences Inc. (Gyeonggi, Korea). This thrombin DNA aptamer (Apt-29; 5'-AGT CCG TGG TAG GGC AGG TTG GGG TGA CT-3') is 29 nucleotides (nt) in length and has high affinity (K d ~ 0.5 nM) [16] for the heparin-binding site of human thrombin [17]. A 5-carboxytetramethylrhodamine (5-TAMRA) -labeled random 29 nt DNA aptamer was purchased from Integrated DNA Technologies Inc. (Coralville, IA, USA).…”

Section: Experimental 21 Reagents and Materials Acrylamide Nn'-mementioning

confidence: 99%

Microchip Electrophoresis Utilizing <i>In Situ</i> Photopolymerized Thrombin-Immobilized Preconcentrator Gels for Specific Entrapment and Analysis of Thrombin Aptamers

et al. 2021

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Section: Experimental 21 Reagents and Materials Acrylamide Nn'-mementioning

confidence: 99%

Microchip Electrophoresis Utilizing <i>In Situ</i> Photopolymerized Thrombin-Immobilized Preconcentrator Gels for Specific Entrapment and Analysis of Thrombin Aptamers

et al. 2021

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“…The development of the aptamer‐based ACE method requires the optimization of several parameters: (i) composition of the BGE, (ii) ion presence (Mg 2+ and K + are important for the aptamer's activity and stability), and (iii) pH and temperature . These methods were used for the detection of several proteins (thrombin, human neutrophil elastase, HIV‐1 reverse transcriptase) , hormones , immunoglobulin E , alkaloids or tumor markers , and even small molecules. Moreover, several research groups used the aptamer‐based ACE technique with nontraditional aptamers for binding studies.…”

Section: Aptamer‐based Affinity Cementioning

confidence: 99%

“…For example, G‐protein‐BODIPY FL GTPγS, hairpin‐structured DNA‐peptide nucleic acid, concanavalin A‐monosaccharides, dorzolamide‐human carbonicanhydrase II, and tubulin‐anti‐mitotic compounds pairs were examined . Other examples of the aptamer‐based ACE applications are summarized in Table .…”

Section: Aptamer‐based Affinity Cementioning

confidence: 99%

Capillary electrophoresis–based immunoassay and aptamer assay: A review

2020

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Capillary electrophoresis-based immunoassay and aptamer assay: A reviewOver the last two decades, the group of techniques called affinity probe CE has been widely used for the detection and the determination of several types of biomolecules with high sensitivity. These techniques combine the low sample consumption and high separation power of CE with the selectivity of the probe to the target molecule. The assays can be defined according to the type of probe used: CE immunoassays, with an antibody as the probe, or aptamer-based CE, with an aptamer as the probe. Immunoassays are generally divided into homogeneous and heterogeneous groups, and homogeneous variant can be further performed in competitive or noncompetitive formats. Interacting partners are free in solution at homogeneous assay, as opposed to heterogeneous analyses, where one of them is immobilized onto a solid support. Highly sensitive fluorescence, chemiluminescence or electrochemical detections were typically used in this type of study. The use of the aptamers as probes has several advantages over antibodies such as shorter generation time, higher thermal stability, lower price, and lower variability. The aptamer-based CE technique was in practice utilized for the determination of proteins in biological fluids and environmentally or clinically important small molecules. Both techniques were also transferred to microchip. This review is focused on theoretical principles of these techniques and a summary of their applications in research.Abbreviations: Ag, antigen; CEIA, CE immunoassay; MCE, microchip CE; SELEX, systematic evolution of ligands by exponential enrichment and they are produced by the immune system of living organisms. Ag that start the immunological response are also called immunogens. These Ab-Ag complexes are stabilized by dipole-dipole, van der Waals or hydrophobic interactions and hydrogen bonds [5,6].The most often used immunoassays are ELISA with Ab or Ag immobilized onto solid support (plates, glass fibers, or plastic tubes), immunofluorescence assay, fluorescence polarization immunoassay, luminescence immunoassay, microparticle enzyme immunoassay, and biosensors. The major disadvantages of immunoassays are the tedious process, time-consuming reactions, and problems with reproducibility and nonspecific interactions [6].CE is a powerful analytical method for the separation of a wide range of molecules. The coupling of CE and immunoassay in the new CE immunoassay (CEIA) technique provided both separation power and specificity of interactions. Other advantages are the possibility of multianalyte analyses, automation, low sample consumption, high speed, flexibility, a lower amount of false-positive results, and also the possibility of connection with highly sensitive detectors [5].In CEIA, the Fab fragments of Ab containing Ag-specific regions are more often used than the whole monoclonal and Color online: See article online to view Figs. 4 and 7 in color.

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“…Consequently, the electrophoretic mobility change is significantly improved, enabling the CE separation of immunocomplex and free aptamers much more easily. Hao et al described a non-competitive CE-LIF-based method to study human thrombin, an essential protein related to the blood coagulation pathway, and successfully detected thrombin in human serum at 0.2 nM using dye-labeled nuclease resistance aptamer Toggle-25 (Figure 2) [66]. Prior to their work, Song et al, reported an affinity probe capillary electrophoresis/laser-induced fluorescence polarization (APCE/LIFP) and achieved the detection of thrombin at low LOD at sub nanomoles/liter [67].…”

Section: Ce-lif-based Immunoassaysmentioning

confidence: 99%

“…( B ) The relationship between the peak area of complex and the concentrations of thrombin. Reproduced under permission of [66].…”

Section: Figurementioning

confidence: 99%

Application of Capillary Electrophoresis with Laser-Induced Fluorescence to Immunoassays and Enzyme Assays

Nguyen

Kang

2019

Molecules

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Capillary electrophoresis using laser-induced fluorescence detection (CE-LIF) is one of the most sensitive separation tools among electrical separation methods. The use of CE-LIF in immunoassays and enzyme assays has gained a reputation in recent years for its high detection sensitivity, short analysis time, and accurate quantification. Immunoassays are bioassay platforms that rely on binding reactions between an antigen (analyte) and a specific antibody. Enzyme assays measure enzymatic activity through quantitative analysis of substrates and products by the reaction of enzymes in purified enzyme or cell systems. These two category analyses play an important role in the context of biopharmaceutical analysis, clinical therapy, drug discovery, and diagnosis analysis. This review discusses the expanding portfolio of immune and enzyme assays using CE-LIF and focuses on the advantages and disadvantages of these methods over the ten years of existing technology since 2008.

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Affinity capillary electrophoresis with laser induced fluorescence detection for thrombin analysis using nuclease-resistant RNA aptamers

Cited by 14 publications

References 33 publications

Microchip Electrophoresis Utilizing <i>In Situ</i> Photopolymerized Thrombin-Immobilized Preconcentrator Gels for Specific Entrapment and Analysis of Thrombin Aptamers

Microchip Electrophoresis Utilizing <i>In Situ</i> Photopolymerized Thrombin-Immobilized Preconcentrator Gels for Specific Entrapment and Analysis of Thrombin Aptamers

Capillary electrophoresis–based immunoassay and aptamer assay: A review

Application of Capillary Electrophoresis with Laser-Induced Fluorescence to Immunoassays and Enzyme Assays

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