{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,9]],"date-time":"2026-10-09T17:53:26Z","timestamp":1791568406469,"version":"4.3.4"},"reference-count":38,"publisher":"MDPI AG","issue":"2","license":[{"start":{"date-parts":[[2025,1,27]],"date-time":"2025-01-27T00:00:00Z","timestamp":1737936000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Intravacc","award":["COMET"],"award-info":[{"award-number":["COMET"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Pathogens"],"abstract":"<jats:p>Hand, foot, and mouth disease (HFMD) represents an emerging health concern whose main causative agents are Coxsackievirus A6 (CVA6) and enterovirus A71 (EV71). The lack of a CVA6 vaccine and the rise of new HFMD-causing strains due to the containment of established HFMD-causing viruses necessitates the search for alternative vaccine technologies, including virus-like particle (VLP) vaccine candidates. While studies have demonstrated that production of enterovirus-like particles in various organisms can be achieved by expression of the viral P1 structural proteins and the 3CD protease, optimization based on the interplay between the three most commonly altered infection parameters (multiplicity of infection (MOI), viable cell density at the time of infection (VCD), and the infection period) is often not investigated. To address this challenge, we have performed Design of Experiments (DoE) to optimize the production of CVA6 and EV71 VLPs. Our results indicate that CVA6 VLP production peaks at high MOI, high VCD, and long infection periods. Our subsequent downstream purification processes yielded 38 mg and 158 mg of purified CVA6 and EV71 VLPs from 1 L crude harvest, respectively. This translates into thousands of potential vaccine doses and highlights the economic potential of enterovirus-like particles for vaccine purposes.<\/jats:p>","DOI":"10.3390\/pathogens14020118","type":"journal-article","created":{"date-parts":[[2025,1,27]],"date-time":"2025-01-27T06:39:51Z","timestamp":1737959991000},"page":"118","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":4,"title":["Optimization of Enterovirus-like Particle Production and Purification Using Design of Experiments"],"prefix":"10.3390","volume":"14","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-7245-3031","authenticated-orcid":false,"given":"Louis","family":"Kuijpers","sequence":"first","affiliation":[{"name":"Department of Bionanoscience, Delft University of Technology, Van der Maasweg 9, 2629 HZ Delft, The Netherlands"},{"name":"Intravacc B.V., Antonie van Leeuwenhoeklaan 9, 3721 MA Bilthoven, The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6170-9051","authenticated-orcid":false,"given":"Wouter J. P.","family":"van den Braak","sequence":"additional","affiliation":[{"name":"Intravacc B.V., Antonie van Leeuwenhoeklaan 9, 3721 MA Bilthoven, The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0005-0303-5553","authenticated-orcid":false,"given":"Abbas","family":"Freydoonian","sequence":"additional","affiliation":[{"name":"Intravacc B.V., Antonie van Leeuwenhoeklaan 9, 3721 MA Bilthoven, The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4029-0973","authenticated-orcid":false,"given":"Nynke H.","family":"Dekker","sequence":"additional","affiliation":[{"name":"Department of Bionanoscience, Delft University of Technology, Van der Maasweg 9, 2629 HZ Delft, The Netherlands"},{"name":"Department of Physics and Kavli Institute of Nanoscience Discovery, University of Oxford, Oxford OX1 3QU, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Leo A.","family":"van der Pol","sequence":"additional","affiliation":[{"name":"Intravacc B.V., Antonie van Leeuwenhoeklaan 9, 3721 MA Bilthoven, The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,1,27]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"500","DOI":"10.1136\/jech-2014-203836","article-title":"Enterovirus 71 related severe hand, foot and mouth disease outbreaks in South-East Asia: Current situation and ongoing challenges","volume":"68","author":"Sabanathan","year":"2014","journal-title":"J. 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