The circular and linear forms of viroid RNAs can be separated by two-dimensional polyacrylamide gel electrophoresis (PAGE) based on the selective delay in mobility that circular RNAs experience under ...denaturing conditions. First PAGE separates RNA preparations from viroid-infected plants, and the whole lane from this first gel is next perpendicularly loaded on top of a second gel. Separation continues under new conditions that differ in the degree of denaturation from the first. The result is a two-dimensional separation of the RNAs in which circular and linear molecules are distributed in two parallel diagonals.
This current review article focuses on recent contributions to on-site forensic investigations. Portable and potentially portable methods are presented and critically discussed about (bio)chemical ...trace analysis and studies performed outside the controlled laboratory environment to rapidly help in crime scene inquiries or forensic intelligence purposes. A wide range of approaches including electrochemical sensors, microchip electrophoresis, ambient ionization on portable mass spectrometers, handheld Raman and NIR instruments as well as and point-of-need devices, like paper-based platforms, for in-field analysis of latent evidences, controlled substances, drug screening, hazards, and others to assist in law enforcements and solving crime more efficiently are highlighted. The covered examples have successfully demonstrated the huge potential of portable devices for on-site applications. Future investigations should consider analytical validation to compete equality and even replace current gold standard methods.
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•Electrochemical sensors offer good sensitivity for abuse drugs and explosives.•Paper-based devices have revealed desirable performance for point-of-care testing.•NIR and RAMAN instruments have allowed fast screening at the point-of-need.•Portable MS instruments have exhibited good performance for on-site forensic applications.•Electrophoresis chips have provided excellent ability for STR genotyping.
Drug binding to plasma proteins influences processes such as liberation, adsorption, disposition, metabolism, and elimination of drugs, which are thus one of the key steps of a new drug development. ...As a result, the characterization of drug–protein interactions is an essential part of these time‐ and money‐consuming processes. It is important to determine not only the binding strength and the stoichiometry of interaction, but also the binding site of a drug on a protein molecule, because two drugs with the same binding site can mutually affect free drug concentration. Capillary electrophoresis‐frontal analysis with mobility shift affinity capillary electrophoresis is one of the most used affinity capillary electrophoresis methods for the characterization of these interactions. In this study, a well‐known sensitivity problem of most capillary electrophoresis‐frontal analyses using ultraviolet detection is solved by its combination with contactless conductivity detection, which provided sixfold lower limits of quantitation and detection. Binding parameters of the human serum albumin‐salicylic acid model affinity pair were evaluated by this newly developed approach and by the classical approach with ultraviolet detection primarily used for their mutual comparison. The results of both approaches agreed well and are also in agreement with literature data obtained using different techniques.
SDS gel electrophoresis is a commonly used approach for monitoring purity and apparent molecular mass (Mr) of proteins, especially in the field of quality control of biopharmaceutical proteins. The ...technological installation of CE‐SDS as the replacement of the slab gel technique (SDS‐PAGE) is still in progress, leading to a continuous improvement of CE‐SDS instruments. Various CE‐SDS instruments, namely Maurice (CE‐SDS/CE‐SDS PLUS) and Wes by ProteinSimple as well as the microchip gel electrophoresis system LabChip® GXII Touch™ HT by PerkinElmer were tested for precision and repeatability compared to SDS‐PAGE (Bio‐Rad). For assessing these quality control parameters, standard model proteins with minor post‐translational modifications were used. Overall, it can be concluded that the CE‐SDS‐based methods are similar to SDS‐PAGE with respect to these parameters. Quality characteristics of test systems gain more significance by testing proteins that do not behave like model proteins. Therefore, glycosylated proteins were analyzed to comparatively investigate the influence of glycosylation on Mr determination in the different instruments. In some cases, high deviations were found both among the methods and with regard to reference values. This article provides possible explanations for these findings.
Methods for studying interactions between glycosaminoglycans (GAGs) and proteins have assumed considerable significance as their biological importance increases. Capillary electrophoresis (CE) is a ...powerful method to study these interactions due to its speed, high efficiency, and low sample/reagent consumption. In addition, CE works effectively under a wide range of physiologically relevant conditions. This chapter presents the state of the art on CE methods for studying GAG-protein interactions including affinity capillary electrophoresis (ACE), capillary zone electrophoresis (CZE), frontal analysis (FA)/frontal analysis continuous capillary electrophoresis (FACCE), and capillary electrokinetic chromatography (CEC) with detailed experimental protocols for ACE and CZE methods.
This review gives a wide overview of recent advances and applications of capillary electrophoresis and microchip capillary electrophoresis methods in the fields of proteomics and peptidomics in the ...period from mid‐2018 up to the end of 2022. The methodological topics covering sample preparation and concentration techniques, hyphenation of capillary electrophoresis methods with mass spectrometry, and multidimensional separations by on‐line or off‐line coupled different capillary electrophoresis and liquid chromatography techniques are described and new developments in both bottom‐up and top‐down approaches in proteomics are presented. In addition, various applications of capillary electrophoresis methods in proteomic and peptidomic studies are demonstrated. They include monitoring of protein posttranslational modifications and applications in biological and biochemical research, clinical peptidomics and proteomics, and food analysis.
•Recent progresses in the chiral separation of amino acids have been discussed.•Different developments in electrodriven separation techniques have been examined.•Enantiorecognition of amino acids by ...using chiral selectors has been explored.
This review highlights recent progresses in the chiral recognition and separation of amino acid enantiomers obtained by capillary electromigration techniques, using different chiral selectors and especially cyclodextrins, covering the literature published from January 2010 to March 2014. Sections are dedicated to the use of derivatization reagents and to the possibility to enantioseparate underivatized amino acids by using either ligand exchange capillary electrophoresis (LECE) and capillary electrophoresis (CE) coupled on line with mass spectrometry. A short insight on frontier nanomaterials is also given.
The temperature-dependent migration of molecular weight protein size standards and several biotherapeutic proteins were studied in sodium dodecyl sulfate capillary gel electrophoresis (SDS-CGE) in ...the interval from 15 to 60 °C using borate cross-linked dextran sieving matrix. Arrhenius plots were generated to calculate the respective activation energy values for the various solute molecules. SDS-CGE analysis of the biotherapeutic protein test mixture revealed no correlation between the activation energy requirement of the different species and their molecular weights, emphasizing the importance of separation temperature optimization to obtain high resolution between the solute molecules of interest. In contrast, the molecular weight protein size ladder ranging from 10 to 225 kDa, built from the same polypeptide blocks with no post-translational and other modifications, showed predictable activation energy requirement. The electrophoretic mobility of the SDS–protein complexes was found to be the function of the reciprocal sixth root of the molecular weight (M w –1/6), implying cylindrical conformation.
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