ELECTROPHORESIS最新文献

{"title":"Enhanced Error Suppression for Accurate Detection of Low-Frequency Variants","authors":"Huimin Chen,&nbsp;Fei Yu,&nbsp;Debin Lu,&nbsp;Shiyue Huang,&nbsp;Songrui Liu,&nbsp;Boseng Zhang,&nbsp;Kunxian Shu,&nbsp;Dan Pu","doi":"10.1002/elps.202400202","DOIUrl":"10.1002/elps.202400202","url":null,"abstract":"<div>\u0000 \u0000 <p>The identification of low-frequency variants remains challenging due to the inevitable high error rates of next-generation sequencing (NGS). Numerous promising strategies employ unique molecular identifiers (UMIs) for error suppression. However, their efficiency depends highly on redundant sequencing and quality control, leading to tremendous read waste and cost inefficiency. Here, we describe a novel approach, enhanced error suppression strategy (EES), that addresses these challenges by (1) optimizing data utilization and reducing read waste by utilizing single-read correction that reserves abundant single reads that complement other single reads or single-strand consensus sequences (SSCSs), and (2) effectively enhancing the accuracy of NGS by employing Bayes’ theorem. EES significantly improves variant detection accuracy, achieving a background error rate of less than 4.4 × 10⁻⁵ per base pair. Additionally, the data utilization rate is dramatically increased, with a 22.9-fold enhancement in duplex consensus sequence (DCS) recovery compared to traditional methodologies. Furthermore, EES demonstrates superior error suppression performance across various base substitutions. In conclusion, EES represents a significant advancement in detecting low-frequency variants by improving data utilization and reducing sequencing errors. It potentially enhances the sensitivity and accuracy of NGS applications, proving highly valuable in clinical and research contexts where precise variant detection is critical.</p>\u0000 </div>","PeriodicalId":11596,"journal":{"name":"ELECTROPHORESIS","volume":"46 1-2","pages":"65-75"},"PeriodicalIF":3.0,"publicationDate":"2024-12-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142827711","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Fundamentals of Viscoelastic Particle Focusing in Suddenly Expanded Microfluidic Channels","authors":"Dalin Chen,&nbsp;Di Huang,&nbsp;Zhixian Zhu,&nbsp;Qiang Huang,&nbsp;Zhonghua Ni,&nbsp;Yiduo Wang,&nbsp;Nan Xiang","doi":"10.1002/elps.202400225","DOIUrl":"10.1002/elps.202400225","url":null,"abstract":"<div>\u0000 \u0000 <p>Particles focusing in suddenly expanded microfluidic channels with viscoelastic biofluids have attracted increased interests due to the potential biomedical applications. In this work, three types of suddenly expanded channels were designed and employed for experiments. These suddenly expanded channels contained a certain number of triangular expansion structures on the sides of the straight channel to investigate the effect of secondary flow on particle viscoelastic focusing and we investigated particle focusing behavior in those three channels with 0.1% hyaluronic acid (HA) solution. The effects of various suddenly expanded channel structures on particle focusing were discussed. The results indicated that the viscoelastic focusing effect was enhanced by using the sudden expansion structures. Consequently, particles with multi-peak distributions were successfully focused into a focusing stream by using the secondary flow induced by the suddenly expanded structure. Finally, the viscoelastic focusing of leukocytes was explored using suddenly expanded channels. This work offers a deep understanding on the fundamentals of particle focusing in suddenly expanded channels and the results will enable the design of particles focusing microfluidic devices with suddenly expanded channels for critical applications in biomedicine.</p>\u0000 </div>","PeriodicalId":11596,"journal":{"name":"ELECTROPHORESIS","volume":"46 3-4","pages":"183-191"},"PeriodicalIF":3.0,"publicationDate":"2024-12-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142827712","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Electrokinetic Power-Series Solution in Narrow Cylindrical Capillaries for All Zeta Potentials.","authors":"Sam Khalifa, Arturo Villegas, Francisco J Diez","doi":"10.1002/elps.202400183","DOIUrl":"https://doi.org/10.1002/elps.202400183","url":null,"abstract":"<p><p>Work from Rice and Whitehead showed the results of electrokinetic flow in a capillary tube under the assumption of low zeta potential <math><semantics><mo><</mo> <annotation>$<$</annotation></semantics> </math> 25 mV, limiting the approximation's usability. Further research conducted by Philip and Wooding provided an alternative solution that assumes high zeta potentials <math><semantics><mo>></mo> <annotation>$>$</annotation></semantics> </math> 25 mV and relies on Rice and Whitehead's solution for lower ranges. However, this solution is presented as a piecewise function, where the functions change based on the zeta potential and the <math> <semantics><mrow><mi>κ</mi> <mi>a</mi></mrow> <annotation>$kappa a$</annotation></semantics> </math> parameter, introducing infinite values for the zeta potential and discontinuities in the derived functions. This paper aims to provide a singular equation solution to the full Poisson-Boltzmann equation for a long cylindrical capillary for all zeta potentials. This solution is a single, continuous, and finite function that produces exact results instead of approximations for all ranges of zeta potential. This exact solution is compared against published approximate solutions for large zeta potentials shown by comparing the large zeta potential approximation with the new exact solution. Important parameters such as volume transport and apparent viscosity were found to have errors of up to 9.76%-57.4%, respectively. The function <math> <semantics><mrow><mi>f</mi> <mo>(</mo> <mi>κ</mi> <mi>a</mi> <mo>,</mo> <msub><mi>ψ</mi> <mn>0</mn></msub> <mo>,</mo> <mi>β</mi> <mo>)</mo></mrow> <annotation>$f(kappa a, psi _0, beta)$</annotation></semantics> </math> has errors of up to 10.5% compared to our full solution.</p>","PeriodicalId":11596,"journal":{"name":"ELECTROPHORESIS","volume":" ","pages":""},"PeriodicalIF":3.0,"publicationDate":"2024-12-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142827601","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"One-Step Focusing of Ampholytes With Electrophoretic Mobilization: Concepts Assessed by Computer Simulation.","authors":"Wolfgang Thormann, Richard A Mosher","doi":"10.1002/elps.202400215","DOIUrl":"https://doi.org/10.1002/elps.202400215","url":null,"abstract":"<p><p>The dynamics of three one-step focusing protocols described in the literature for IEF-MS analyses of proteins are assessed by computer simulation. Focusing of 101 carrier ampholytes (pI range 3.0-11.0) and 9 analytes (pI range 3.9-10.4) with concurrent electrophoretic mobilization in an electroosmosis-free environment is studied via use of the following: (1) an acid as anolyte and catholyte, (2) a plug of a base between the sample and the acid as catholyte, and (3) a base incorporated into the acid-based catholyte. The data reveal that the first approach is of limited applicability and separation efficiency as it provides separated transient foci of acidic ampholytes only. The second system largely extends the applicability range, separation, and sensitivity and can be applied to ampholytes with pIs as high as 10. The same is true with the base incorporated into the catholyte. Dynamic simulation provides for the first time insight into the complexity of these one-step focusing systems with buffer components that are compatible with MS detection. Simulation is an attractive and simple tool for characterization of the involved electrophoretic processes and for assay optimization.</p>","PeriodicalId":11596,"journal":{"name":"ELECTROPHORESIS","volume":" ","pages":""},"PeriodicalIF":3.0,"publicationDate":"2024-12-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142784529","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Quantitative Endogenous Polyamine Analysis via Capillary Electrophoresis/Mass Spectrometry: Characterization and Practical Considerations","authors":"Cameron J. Kaminsky,&nbsp;Jericha Mill,&nbsp;Thomas Raife,&nbsp;Lingjun Li","doi":"10.1002/elps.202400165","DOIUrl":"10.1002/elps.202400165","url":null,"abstract":"<p>Commonly used analytical techniques for polyamine analysis, including derivatization and mixed-mode liquid chromatography (LC), have inherent disadvantages. Capillary electrophoresis (CE) is uniquely suited to analyze small, highly charged molecules because analytes are separated on the basis of their electrophoretic mobility, not polarity or association with a stationary phase. Microfluidic CE–mass spectrometry (mCE–MS) is a relatively recent addition to commercially available CE offerings that streamlines traditional CE–MS interfacing and has the potential to improve upon classic CE challenges to robustness and reproducibility. MS instrument choice and scanning parameters are strongly influenced by a need for high acquisition rate to adequately sample CE peaks. Alternatively, isotachophoresis on loading can be intentionally avoided to produce sufficiently wide peaks. The mCE platform utilized here performed very well in many metrics; a limit of detection (LOD) as low as 0.25 ng/mL was achieved for spermidine, and endogenous spermidine was easily detected in blood with this method. Both of these are challenging tasks for any separation technique and demonstrate a strong use case for the platform. During experimentation, various idiosyncrasies in the commercial CE–MS interface resulted in extensive chip-to-chip variability in both peak shape and LOD, complicating the application to robust absolute quantitation. Practical guidance for similar analyses is provided.</p>","PeriodicalId":11596,"journal":{"name":"ELECTROPHORESIS","volume":"46 5-6","pages":"296-304"},"PeriodicalIF":3.0,"publicationDate":"2024-12-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/elps.202400165","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142784563","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A Low-Cost Microfluidic Device For the On-Line Counting of Microparticle/Bacteria","authors":"Zhen-Yu Xun,&nbsp;Lv Liu,&nbsp;Bai-Chuan Zhang,&nbsp;Liang-Liang Fan,&nbsp;Lu-Wei Zhang,&nbsp;Hong Zhao,&nbsp;Sohail Iqbal,&nbsp;Rana Iqtidar Shakoor,&nbsp;Liang Zhao","doi":"10.1002/elps.202400061","DOIUrl":"10.1002/elps.202400061","url":null,"abstract":"<div>\u0000 \u0000 <p>On-line counting of the microparticle/bacteria in the liquid medium has great potential in the food safety and biomedical fields. A new low-cost microfluidic device is proposed for the on-line counting of the microparticles/bacteria in the liquid medium. The gradually contracted microchannel and the viscoelastic fluid are combined to achieve the efficient elastic focusing of the particle/bacteria, which significantly improves the counting accuracy by aligning all particles/bacteria in a single position at the center of the microchannel. A simple light resistance-based counting method is designed and integrated with the microchannel, where the low-cost elements including the laser pointer, convex lens, diaphragm, and photodiode, are used to build the optical path of detection. The influence of the flow rate, the particle size, the property of fluid, and the channel structure on the focusing of the particle/bacteria are investigated by the experiment, and the counting ability of the integrated microfluidic device is validated by using different-sized microparticles and the <i>bifidobacterium</i>. With its simple structure, low cost, easy operation, and high efficiency, this microfluidic device is suitable for the commercial applications, such as the on-line counting of the plastic microparticle in water or the colony-forming units (CFU) of bacteria in food.</p>\u0000 </div>","PeriodicalId":11596,"journal":{"name":"ELECTROPHORESIS","volume":"45 23-24","pages":"2182-2190"},"PeriodicalIF":3.0,"publicationDate":"2024-12-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142767232","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Electromigration of Charged Analytes Through Immiscible Fluids in Multiphasic Electrophoresis","authors":"Md Nazibul Islam,&nbsp;Yang Liu,&nbsp;Amy E. Herr","doi":"10.1002/elps.202400192","DOIUrl":"10.1002/elps.202400192","url":null,"abstract":"<p>Multiphasic buffer systems have been of greatest interest in electrophoresis and liquid–liquid electrotransfer; this study extends that foundation by exploring the interplay of the geometric and viscous properties of an interleaving oil layer on the electrotransfer of a charged analyte from an aqueous solution into a hydrogel. We utilized finite element analysis to examine two complementary configurations: one being electrotransfer of a charged analyte (protein) in an aqueous phase into a surrounding hydrogel layer and another being electrotransfer of the protein from that originating aqueous phase—through an interleaving oil layer of predetermined viscosity and thickness—and into a surrounding hydrogel layer. Results indicate that the presence of an oil layer leads to increased skew of the injected peak. To explain this difference in injection dispersion, we utilize Probstein's framework and compare the Péclet (<i>Pe</i>) number with the ratio between length scales characteristic to the axial and radial dispersion, respectively. The formulation assigns electrotransfer conditions into six different dispersion regimes. We show that the presence or absence of an interleaving oil layer moves the observed peak dispersion into distinct electrotransfer regimes; the presence of an oil layer augments the electrophoretic mobility mismatch among the different phases, resulting in a five-fold increase in <i>Pe</i> and a six-fold increase in the ratio between the axial to radial dispersion characteristic lengths. We further show that oil viscosity significantly influences resultant injection dispersion. A decrease in oil-layer viscosity from 0.08 to 0.02 Pa s results in a &gt;100% decrease in injection dispersion. Our theoretical predictions were experimentally validated by comparing the electrotransfer regimes of three different mineral oil samples. We show that lowering the oil viscosity to 0.0039 Pa s results in an injection regime similar to that of the absence of an oil layer. Understanding of the impact of electrotransfer of charged species across multiple immiscible fluid layers on peak dispersion informs the design of multiphasic electrophoresis systems.</p>","PeriodicalId":11596,"journal":{"name":"ELECTROPHORESIS","volume":"46 1-2","pages":"13-21"},"PeriodicalIF":3.0,"publicationDate":"2024-12-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11773303/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142767247","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Determination of the Effective Charge Numbers and Ionic Mobilities of Single Isomer and Randomly Highly Sulfated Cyclodextrins by Capillary Isotachophoresis and Zone Electrophoresis.","authors":"Veronika Šolínová, Dušan Koval, Václav Kašička","doi":"10.1002/elps.202400207","DOIUrl":"https://doi.org/10.1002/elps.202400207","url":null,"abstract":"<p><p>Sulfated cyclodextrins (CDs) are multiply negatively charged molecules widely used as chiral selectors in capillary electrophoresis (CE). In some of their applications, the effective charge numbers of their molecules were observed to be lower than the numbers of the attached sulfated groups due to strong binding of counterions. However, degree of reduction of the theoretical charge was not quantified. For that reason, in this study, capillary isotachophoresis (CITP) and capillary zone electrophoresis (CZE) were applied for the determination of the effective charge numbers and actual ionic mobilities of two kinds of sulfated CDs: single isomer sulfated α-, β-, and γ-CDs (SI-CDs) and randomly highly sulfated α-, β-, and γ-CDs (HS-CDs). The effective charge numbers of the SI-CDs and HS-CDs were determined from the length of their ITP zones, the ionic mobilities determined by CZE, and molar concentrations of their solutions applied for CITP analysis, and from the same parameters of reference compounds, formic acid for SI-CDs and dichloroacetic acid for HS-CDs. The determined effective charge numbers of the SI-CDs were equal to or only slightly lower than the numbers of sulfate groups in their molecules but the effective charge numbers of randomly HS-CDs were significantly (22.2%-27.8%) reduced as compared to the average numbers of sulfate groups in their molecules. In accordance with a lower number of sulfate groups in SI-CDs than in HS-CDs, the absolute values of the actual ionic mobilities of SI-CDs (35.5-37.5) × 10^-9 m² V^-1 s^-1 were lower than those of HS-CDs (43.5-44.1) × 10^-9 m² V^-1 s^-1.</p>","PeriodicalId":11596,"journal":{"name":"ELECTROPHORESIS","volume":" ","pages":""},"PeriodicalIF":3.0,"publicationDate":"2024-11-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142738718","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Enhancing Doxorubicin Detection: Multiphase Electroextraction for Efficient and Affordable UHPLC-DAD Analysis in Saliva.","authors":"Denise V M Sousa, Fabiano V Pereira, Ricardo M Orlando","doi":"10.1002/elps.202400094","DOIUrl":"https://doi.org/10.1002/elps.202400094","url":null,"abstract":"<p><p>Attesting optimal drug concentrations in biological fluids is crucial to ensure precise dosage adjustment, to guarantee therapy adherence, and to manage side effects in chemotherapy. Accurate drug determination relies on liquid chromatography and advanced detectors, with sample preparation playing a pivotal role, especially in complex matrices such as biological fluids. This study introduces a multiphase electroextraction (MPEE) of doxorubicin (DOX) in saliva by utilizing a paper point, followed by ultra-high-performance liquid chromatography coupled to diode array detection. The extraction time and electric potential were carried out by using the Doehlert optimization approach, whereas the desorption solvent was fine-tuned through the centroid-simplex experimental design. After optimization, DOX and the internal standard were extracted in 35 min, utilizing an applied voltage of 300 V and a multiwell plate capable of simultaneous extraction of 66 samples. The recovery was 87%-101%, with a linear range between 50 and 500 µg L^-1 (R² > 0.999). The intra- and inter-assay coefficients of variation for precision were <10%, and the limit of detection and limit of quantification were 25 and 50 µg L^-¹, respectively. When applied to five different fortified saliva samples, there were no statistically significant differences in the detected concentrations. Although the enrichment factor (0.6) was not as high as expected, the other results confirm that the method obtained is suitable for monitoring DOX in this complex matrix and can contribute to further developments in sample preparation using MPEE approaches.</p>","PeriodicalId":11596,"journal":{"name":"ELECTROPHORESIS","volume":" ","pages":""},"PeriodicalIF":3.0,"publicationDate":"2024-11-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142738719","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"When to Use Rectangular Waveforms in Dielectrophoresis Application to Increase Separation and Sorting Efficiency","authors":"Niklas P. Boldt,&nbsp;Laura Weirauch,&nbsp;Jana M. Späth,&nbsp;Uwe Kerst,&nbsp;Mario Birkholz,&nbsp;Michael Baune,&nbsp;Roland Thewes","doi":"10.1002/elps.202400164","DOIUrl":"10.1002/elps.202400164","url":null,"abstract":"<p>In this study, the influence of using rectangular waveforms is comprehensively investigated on the separation and sorting efficiency of dielectrophoretic (DEP) processes. Besides positive effects on DEP experiments, cases of a diminished force due to rectangular waveforms are investigated and discussed. This investigation encompasses two primary experimental setups. First, microparticle-focusing experiments are carried out using a pair of electrodes within a microfluidic channel. Second, separation experiments are performed using a macroscopic insulator-based dielectrophoretic filter. The study reveals that harmonics of rectangular signals can have a positive impact on separation or sorting efficiency when compared to sinusoidal waveforms, provided that these harmonics contribute to the overall DEP force with the same sign. This positive effect is found to depend on the ratio between the applied fundamental frequency and the cross-over frequency in the Clausius–Mossotti factor. However, violating related derived boundary conditions leads to negative effects and a decrease in the DEP net force.</p>","PeriodicalId":11596,"journal":{"name":"ELECTROPHORESIS","volume":"46 1-2","pages":"104-111"},"PeriodicalIF":3.0,"publicationDate":"2024-11-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11773296/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142738725","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}