Capillary electrophoresis–based immunoassay and aptamer assay: A review

Abstract

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.

Immunoaffinity Capillary Electrophoresis in the Era of Proteoforms, Liquid Biopsy and Preventive Medicine: A Potential Impact in the Diagnosis and Monitoring of Disease Progression

“…In general, affinity capillary electrophoresis is a broad term referring to the separation by capillary electrophoresis of substances that participate in specific or non-specific noncovalent affinity interactions during electrophoresis. The interacting molecules can be found in solution or can be immobilized to a solid support [123,146,[174][175][176][177][178][179][180][191][192][193][194][195][196][197][198][199][200][201][202][203]. Immunoaffinity capillary electrophoresis, is the method referred to when using immobilized affinity ligands or probes to capture one or more analytes found in simple or complex matrices.…”

A Two-Dimensional Affinity Capture and Separation Mini-Platform for the Isolation, Enrichment, and Quantification of Biomarkers and Its Potential Use for Liquid Biopsy

“…One technology in particular is the combination of solid-phase extraction methods, using affinity-based ligands, with CE. Highly selective affinity ligands or adsorbents, such as antibodies, antibody fragments, lectins, aptamers, enzymes, metal-organic framework-based affinity sorbents, and other specially designed ligands are the cornerstone for the isolation and purification of many substances and cellular-subcellular entities found in complex matrices [ 25 , 47 , 54 , 55 , 56 , 57 , 58 , 75 , 81 , 82 , 83 , 84 , 85 , 86 , 87 , 88 , 89 , 90 , 91 , 92 , 93 , 94 , 95 , 96 , 97 , 98 ].…”

“…The high-resolution analytical separation ability of IACE provides the advantage of characterizing and discerning different forms of the same protein (proteoforms) biomarker and drug metabolites, which can improve insights into disease diagnosis, monitor drug efficacy, and shorten the length of clinical trials. Since the introduction of IACE in the early 1990s by our laboratory, Terry Phillips’ laboratory, and others, the technology has increased significantly in popularity, and many applications have been reported using both conventional CE and microchip CE [ 25 , 54 , 55 , 58 , 61 , 62 , 63 , 64 , 66 , 75 , 88 , 89 , 90 , 91 , 92 , 93 , 94 , 95 , 96 , 97 , 98 ]. Furthermore, if IACE is coupled to powerful detectors such as laser-induced fluorescence detectors or mass spectrometers, a significant improvement in limits of detection can been achieved [ 25 , 75 , 99 , 100 , 101 , 102 , 103 , 104 , 105 , 106 , 107 , 108 ].…”

“…Numerous immunoaffinity capillary applications employing various configurations of the ACM device, using either a conventional capillary electrophoresis or a microfluidic chip capillary electrophoresis, have been reported [ 25 , 54 , 55 , 56 , 57 , 58 , 61 , 75 , 88 , 89 , 90 , 91 , 92 , 93 , 94 , 95 , 96 , 97 , 98 ]. Figure 2 depicts the use of an IACE selective affinity capture-separation system for the quantification of brain-derived neurotrophic factor (BDNF), a neuropeptide growth factor obtained from human skin biopsies of atopic inflammatory reactions [ 124 ].…”

Capillary electrophoresis‐based methodology for screening of oligonucleotide aptamers

“…Compared with antibodies, aptamers have several significant advantages: (a) they have high chemical stability and are insensitive to the change in pH value, and can be stored at ambient temperature for a long time; (b) the denaturation of aptamers is highly reversible, that is to say, aptamers can be denatured at high temperature and renatured at low temperature; (c) they are easy to obtain, because they can be screened nonbiologically in vitro, and can be synthesized with high purity when determining the sequence; (d) they can recognize many kinds of targets, including proteins, nucleic acids, cells and various small-molecule compounds; (e) they have high affinity to targets with dissociation constants in the range of 10 −6 -10 −12 M; and (f) they are easy to modify and are suitable for binding with nanomaterials. In view of this, aptamers have been widely used in biomedical, analytical and other fields (Dunn et al, 2017;Groff et al, 2015;Hamula et al, 2011;Kong & Byun, 2013;Nevídalová et al, 2020;Song et al, 2019;Tombelli et al, 2007;Yoshimoto, 2019).…”