{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,15]],"date-time":"2026-09-15T16:04:55Z","timestamp":1789488295309,"version":"build-2803163510"},"reference-count":34,"publisher":"American Chemical Society (ACS)","issue":"36","license":[{"start":{"date-parts":[[2026,8,28]],"date-time":"2026-08-28T00:00:00Z","timestamp":1787875200000},"content-version":"stm-asf","delay-in-days":0,"URL":"https:\/\/doi.org\/10.15223\/policy-029"},{"start":{"date-parts":[[2026,8,28]],"date-time":"2026-08-28T00:00:00Z","timestamp":1787875200000},"content-version":"stm-asf","delay-in-days":0,"URL":"https:\/\/doi.org\/10.15223\/policy-037"},{"start":{"date-parts":[[2026,8,28]],"date-time":"2026-08-28T00:00:00Z","timestamp":1787875200000},"content-version":"stm-asf","delay-in-days":0,"URL":"https:\/\/doi.org\/10.15223\/policy-045"}],"funder":[{"DOI":"10.13039\/100022752","name":"MSD Sharp and Dohme","doi-asserted-by":"publisher","award":["NA"],"award-info":[{"award-number":["NA"]}],"id":[{"id":"10.13039\/100022752","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2026,9,15]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>Host cell residual DNA (rDNA) is a critical process-related impurity in biopharmaceutical production, involving trace amounts of DNA fragments originating from yeast, bacteria, CHO (Chinese hamster ovary), and other host cells. Monitoring both the size and quantity of rDNA is essential because of potential safety risks, including oncogenicity and immunogenicity, in biological products such as vaccines and other protein therapeutics. Regulatory guidelines from the WHO and U.S. FDA recommend limits of less than 10 ng\/dose and below 200 base pairs (bp) for the rDNA content and size in final products. In this work, innovative capillary electrophoresis (CE) technologies that can accurately size rDNA were explored using a single-capillary (BIABooster) CE system and confirmed with a 12-capillary (Femto Pulse) CE system. After optimizing the sample diluent, detection parameters, and hydrodynamic injection volumes, the BIABooster method achieved highly sensitive detection at 200 bp in the linear range of 0.035 ng\/mL to 1 ng\/mL, with an LOD of 0.035 ng\/mL (i.e., 0.0175 ng in 0.5 mL dose, S\/N = 3) and an LOQ of 0.065 ng\/mL (i.e, 0.0325 ng in 0.5 mL dose, S\/N = 10). As a proof of concept, this method was successfully applied to separate rDNA in real biopharmaceutical samples, including yeast-derived in-process intermediate (IPI) samples and CHO-derived final products, showing high sizing accuracy (99.8\u2013104.2%) and robust performance with an intermediate precision of 2.5% RSD from 200 bp to 1000 bp, across preparations, days, analysts, and capillary lots. Compared to conventional methods in rDNA analysis, this method offers advantages including high sensitivity and accuracy, automation, and no need for DNA probe design and amplification steps, resulting in faster method development.<\/jats:p>","DOI":"10.1021\/acs.analchem.6c04185","type":"journal-article","created":{"date-parts":[[2026,9,2]],"date-time":"2026-09-02T05:11:28Z","timestamp":1788325888000},"page":"27043-27052","source":"Crossref","is-referenced-by-count":0,"title":["Sensitive and\nRobust Capillary Electrophoretic Analysis\nof Host Cell Residual DNA in Biopharmaceutical Samples"],"prefix":"10.1021","volume":"98","author":[{"given":"Laura N.","family":"Taylor","sequence":"first","affiliation":[{"name":"Merck & Co., Inc. , , , ,","place":["Rahway, New Jersey, United States, 07065"]},{"name":"West Virginia University , , , ,","place":["Morgantown, West Virginia, United States, 26505"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2156-9963","authenticated-orcid":true,"given":"Paul F.","family":"Gillespie","sequence":"additional","affiliation":[{"name":"Merck & Co., Inc. , , , ,","place":["Rahway, New Jersey, United States, 07065"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Richard R.","family":"Rustandi","sequence":"additional","affiliation":[{"name":"Merck & Co., Inc. , , , ,","place":["Rahway, New Jersey, United States, 07065"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Matthew","family":"Schombs","sequence":"additional","affiliation":[{"name":"Merck & Co., Inc. , , , ,","place":["Rahway, New Jersey, United States, 07065"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6492-6473","authenticated-orcid":true,"given":"Adam T.","family":"Sutton","sequence":"additional","affiliation":[{"name":"Merck & Co., Inc. , , , ,","place":["Rahway, New Jersey, United States, 07065"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1439-9548","authenticated-orcid":true,"given":"Wan","family":"Zhou","sequence":"additional","affiliation":[{"name":"Merck & Co., Inc. , , , ,","place":["Rahway, New Jersey, United States, 07065"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"316","published-online":{"date-parts":[[2026,8,28]]},"reference":[{"issue":"2","key":"2026091511064191500_cit1","doi-asserted-by":"publisher","first-page":"307","DOI":"10.1002\/bit.23343","article-title":"Residual DNA Analysis\nin Biologics Development: Review of Measurement and Quantitation Technologies\nand Future Directions","volume":"109","author":"Wang","year":"2012","journal-title":"Biotechnol. Bioeng."},{"issue":"2","key":"2026091511064191500_cit2","doi-asserted-by":"publisher","first-page":"155","DOI":"10.5731\/pdajpst.2013.00910","article-title":"Establishing\nAcceptable Limits of Residual DNA","volume":"67","author":"Yang","year":"2013","journal-title":"PDA J. Pharm.\nSci. Technol."},{"key":"2026091511064191500_cit3","unstructured":"Guidance for Industry- Characterization and\nQualification of Cell\nSubstrates and Other Biological Materials Used in the Production of\nViral Vaccines for Infectious Disease Indications; https:\/\/www.fda.gov\/media\/78428\/download (accessed 2026\u201305\u201330)."},{"key":"2026091511064191500_cit4","unstructured":"World Health Organization (WHO).\n          .  WHO Expert Committee on Biological Standardization.\nFifty-Sixth Report; WHO Technical Report Series, No. 941.; World Health Organization (WHO Press), Geneva, Switzerland, 2007. https:\/\/iris.who.int\/server\/api\/core\/bitstreams\/81d62ecc-5708-480d-a13d-4a49e2a5f704\/content (accesed May 31, 2026)."},{"issue":"1","key":"2026091511064191500_cit5","doi-asserted-by":"publisher","first-page":"17","DOI":"10.1186\/s12575-019-0105-1","article-title":"Development and Validation\nof Quantitative Real-Time PCR for the\nDetection of Residual CHO Host Cell DNA and Optimization of Sample\nPretreatment Method in Biopharmaceutical Products","volume":"21","author":"Zheng","year":"2019","journal-title":"Biol. Proced. Online"},{"key":"2026091511064191500_cit6","doi-asserted-by":"publisher","first-page":"115577","DOI":"10.1016\/j.ab.2024.115577","article-title":"PicoGreen Assay for\nNucleic Acid Quantification - Applications, Challenges,\nand Solutions","volume":"692","author":"Ban","year":"2024","journal-title":"Anal. Biochem."},{"issue":"4","key":"2026091511064191500_cit7","doi-asserted-by":"publisher","DOI":"10.4155\/fsoa-2017-0115","article-title":"Validation of Quantitative\nPolymerase Chain Reaction with Southern Blot Method for Telomere Length\nAnalysis","volume":"4","author":"Tarik","year":"2018","journal-title":"Future Sci. OA"},{"issue":"7","key":"2026091511064191500_cit8","doi-asserted-by":"publisher","first-page":"997","DOI":"10.3390\/biology12070997","article-title":"Next-Generation Sequencing Technology: Current Trends\nand Advancements","volume":"12","author":"Satam","year":"2023","journal-title":"Biology"},{"key":"2026091511064191500_cit9","doi-asserted-by":"publisher","first-page":"1464","DOI":"10.1021\/acs.analchem.8b00015","article-title":"Capillary Electrophoresis:\nTrends and Recent Advances. Analytical Chemistry","volume":"6","author":"Voeten","year":"2018","journal-title":"American Chemical Society February"},{"key":"2026091511064191500_cit10","first-page":"10.9.1","article-title":"Capillary Electrophoresis\nof DNA","volume-title":"Current Protocols in Nucleic Acid\nChemistry","author":"Smith","year":"2003"},{"issue":"19","key":"2026091511064191500_cit11","doi-asserted-by":"publisher","first-page":"7885","DOI":"10.1021\/ac901681k","article-title":"Fluorescence Polarization\nCombined Capillary Electrophoresis Immunoassay\nfor the Sensitive Detection of Genomic DNA Methylation","volume":"81","author":"Wang","year":"2009","journal-title":"Anal. Chem."},{"key":"2026091511064191500_cit12","doi-asserted-by":"publisher","first-page":"136","DOI":"10.1016\/j.aca.2015.03.009","article-title":"Reversible Phospholipid Nanogels for Deoxyribonucleic\nAcid Fragment Size Determinations up to 1500 Base Pairs and Integrated\nSample Stacking","volume":"880","author":"Durney","year":"2015","journal-title":"Anal. Chim. Acta"},{"key":"2026091511064191500_cit13","doi-asserted-by":"crossref","first-page":"359","DOI":"10.1006\/abio.1995.9986","article-title":"Analysis of Double-Stranded\nDNA by Capillary Electrophoresis\nwith Laser-Induced Fluorescence Detection Using the Monomeric Dye\nSYBR Green I","volume":"231","author":"Skeidsvoll","year":"1995","journal-title":"Anal. Biochem."},{"issue":"1","key":"2026091511064191500_cit14","doi-asserted-by":"publisher","first-page":"195","DOI":"10.1016\/S0021-9673(02)00697-0","article-title":"Effects of Metal\nIons on Concentration\nof DNA in High-Conductivity Media by Capillary Electrophoresis","volume":"966","author":"Huang","year":"2002","journal-title":"J. Chromatogr. A"},{"issue":"11","key":"2026091511064191500_cit15","doi-asserted-by":"publisher","first-page":"1555","DOI":"10.1002\/elps.201200624","article-title":"CGE-Laser\nInduced Fluorescence of Double-Stranded DNA Fragments Using GelGreen\nDye","volume":"34","author":"Valde\u0301s","year":"2013","journal-title":"Electrophoresis"},{"issue":"6","key":"2026091511064191500_cit16","doi-asserted-by":"publisher","first-page":"3766","DOI":"10.1021\/acs.analchem.7b04034","article-title":"BIABooster: Online\nDNA Concentration\nand Size Profiling with a Limit of Detection of 10 Fg\/\u039cL and\nApplication to High-Sensitivity Characterization of Circulating Cell-Free\nDNA","volume":"90","author":"Andriamanampisoa","year":"2018","journal-title":"Anal. Chem."},{"key":"2026091511064191500_cit17","doi-asserted-by":"publisher","first-page":"53","DOI":"10.1007\/978-1-0716-3850-7_2","article-title":"DNA Sizing\nwith pg\/\u03bcL\nSensitivity for Cell-Free DNA Analysis with the \u03bcLAS Technology","volume":"2804","author":"Boutonnet","year":"2024","journal-title":"Methods Mol Biol."},{"key":"2026091511064191500_cit18","unstructured":"Agilent Technologies.\n          \n          Femto Pulse\nSystem. https:\/\/www.agilent.com\/en\/product\/automated-electrophoresis\/femto-pulse-systems\/femto-pulse-system\/femto-pulse-system-365750 (accessed 2025\u201310\u201316)."},{"key":"2026091511064191500_cit19","unstructured":"Agilent Technologies.\n          \n          Agilent FP-1101 Ultra\nSensitivity NGS Kit; Agilent Ultra Sensitivity NGS Kit Quick Guide\nfor the Femto Pulse System, 2025. SD-AT000143\nEdition 03\/25. https:\/\/www.agilent.com\/cs\/library\/usermanuals\/public\/quick-guide-fp-1101-ultra-sensitivity-ngs-kit-SD-AT000143.pdf."},{"key":"2026091511064191500_cit20","unstructured":"Adelis Advacned Separative Technologies.\n          \n          DNA 1K Kit BIABooster CE Pack, 2025. https:\/\/www.adelis-tech.com\/wp-content\/uploads\/2024\/04\/ProductNote_DNA1K_CE.pdf (accessed 2025\u201310\u201316)."},{"issue":"24","key":"2026091511064191500_cit21","doi-asserted-by":"publisher","first-page":"9263","DOI":"10.1021\/acs.analchem.3c00998","article-title":"Size and Concentration\nof Cell-Free DNA Measured Directly from Blood Plasma, without Prior\nDNA Extraction","volume":"95","author":"Boutonnet","year":"2023","journal-title":"Anal. Chem."},{"key":"2026091511064191500_cit22","doi-asserted-by":"publisher","first-page":"6923","DOI":"10.1007\/s00216-015-8703-5","article-title":"Capillary Electrophoresis Applied\nto DNA: Determining and Harnessing Sequence and Structure to Advance\nBioanalyses (2009\u20132014)","volume":"407","author":"Durney","year":"2015","journal-title":"Anal. Bioanal.\nChem."},{"issue":"10","key":"2026091511064191500_cit23","doi-asserted-by":"publisher","first-page":"2634","DOI":"10.1021\/ja01265a018","article-title":"The Influence\nof Ionic Strength on the Quenching of\nFluorescence in Aqueous Solutions","volume":"61","author":"Stoughton","year":"1939","journal-title":"J. Am. Chem.\nSoc."},{"key":"2026091511064191500_cit24","doi-asserted-by":"publisher","first-page":"119","DOI":"10.1016\/S0021-9673(00)00812-8","article-title":"Sample Stacking of Cationic and Anionic Analytes in\nCapillary Electrophoresis","volume":"902","author":"Quirino","year":"2000","journal-title":"J. Chromatogr. A"},{"key":"2026091511064191500_cit25","doi-asserted-by":"publisher","first-page":"15","DOI":"10.1186\/s41935-024-00389-y","article-title":"An Overview\nof DNA Degradation and\nIts Implications in Forensic Caseworks","volume":"14","author":"Bhoyar","year":"2024","journal-title":"Egyptian\nJournal of Forensic Sciences"},{"issue":"4","key":"2026091511064191500_cit26","doi-asserted-by":"publisher","first-page":"242","DOI":"10.3390\/gels10040242","article-title":"Quantitative\nAnalysis of Genomic\nDNA Degradation of E. Coli Using Automated Gel Electrophoresis under\nVarious Levels of Microwave Exposure","volume":"10","author":"Pandey","year":"2024","journal-title":"Gels"},{"key":"2026091511064191500_cit27","doi-asserted-by":"publisher","first-page":"228","DOI":"10.1006\/abio.1997.2177","article-title":"Characterization\nof PicoGreen Reagent and Development of a Fluorescence-Based Solution\nAssay for Double-Stranded DNA Quantitation","volume":"249","author":"Singer","year":"1997","journal-title":"Analytical Biochemistry"},{"key":"2026091511064191500_cit28","doi-asserted-by":"publisher","first-page":"127","DOI":"10.1080\/bih.76.3.127.132","article-title":"Staining\nnucleic acids and proteins in electrophoresis gels","volume":"76","author":"Williams","year":"2001","journal-title":"Biotechnic & Histochemistry"},{"key":"2026091511064191500_cit29","doi-asserted-by":"publisher","first-page":"108","DOI":"10.3389\/fmicb.2017.00108","article-title":"A Basic Guide to Real\nTime PCR in Microbial Diagnostics:\nDefinitions, Parameters, and Everything","volume":"8","author":"Kralik","year":"2017","journal-title":"Frontiers\nin Microbiology"},{"issue":"1","key":"2026091511064191500_cit30","doi-asserted-by":"publisher","first-page":"74","DOI":"10.1186\/1471-2164-11-74","article-title":"Automated Design\nof Genomic Southern Blot Probes","volume":"11","author":"Croning","year":"2010","journal-title":"BMC Genomics"},{"issue":"4","key":"2026091511064191500_cit31","doi-asserted-by":"publisher","first-page":"1999","DOI":"10.3390\/s23041999","article-title":"Uncertainty Estimation\nfor Quantitative Agarose Gel Electrophoresis of Nucleic Acids","volume":"23","author":"Semenov","year":"2023","journal-title":"Sensors"},{"key":"2026091511064191500_cit32","doi-asserted-by":"publisher","first-page":"137625","DOI":"10.1016\/j.snb.2025.137625","article-title":"Integrated Microfluidic Device of\nDNA Extraction, Recombinase Polymerase Amplification and Micro-Capillary\nElectrophoresis for Sample-to-Answer Detection of Salmonella Typhimurium","volume":"435","author":"Nguyen","year":"2025","journal-title":"Sens. Actuators B Chem."},{"issue":"51","key":"2026091511064191500_cit33","doi-asserted-by":"publisher","first-page":"20267","DOI":"10.1021\/acs.analchem.4c04822","article-title":"Integration of Immunosyringe\nSensors with Bar-Chart Chips for Prick-and-Read Testing of Prostate\nSpecific Antigen","volume":"96","author":"Li","year":"2024","journal-title":"Anal. Chem."},{"issue":"14","key":"2026091511064191500_cit34","doi-asserted-by":"publisher","first-page":"5625","DOI":"10.1021\/acs.analchem.4c00255","article-title":"Microfluidic\nImmunosensing Platform Based on a Rolling Circle Amplification-Assisted\nDNA Dendrimer Probe for Portable and Sensitive Detection of Allergen-Specific\nIgE","volume":"96","author":"Chen","year":"2024","journal-title":"Anal. Chem."}],"container-title":["Analytical Chemistry"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/pubs.acs.org\/doi\/pdf\/10.1021\/acs.analchem.6c04185","content-type":"application\/pdf","content-version":"vor","intended-application":"unspecified"},{"URL":"https:\/\/pubs.acs.org\/ancham\/article-pdf\/98\/36\/27043\/67623944\/acs.analchem.6c04185.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/pubs.acs.org\/ancham\/article-pdf\/98\/36\/27043\/67623944\/acs.analchem.6c04185.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,9,15]],"date-time":"2026-09-15T15:26:19Z","timestamp":1789485979000},"score":1,"resource":{"primary":{"URL":"https:\/\/pubs.acs.org\/ancham\/article\/98\/36\/27043\/5351094\/Sensitive-and-Robust-Capillary-Electrophoretic"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,8,28]]},"references-count":34,"journal-issue":{"issue":"36","published-online":{"date-parts":[[2026,8,28]]},"published-print":{"date-parts":[[2026,9,15]]}},"URL":"https:\/\/doi.org\/10.1021\/acs.analchem.6c04185","relation":{},"ISSN":["0003-2700","1520-6882"],"issn-type":[{"value":"0003-2700","type":"print"},{"value":"1520-6882","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,8,28]]}}}