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Definitive diagnosis currently relies on post-mortem histopathological examination of the central nervous system. Among emerging diagnostic tools,\n                      <jats:italic toggle=\"yes\">in vitro<\/jats:italic>\n                      amplification techniques such as Real-Time Quaking-Induced Conversion (RT-QuIC) have demonstrated high sensitivity, specificity, and speed, although certain prion strains remain difficult to amplify. Here, we evaluate a novel recombinant substrate for RT-QuIC: human PrP E219K, a naturally occurring polymorphism in which lysine substitutes glutamic acid at codon 219. Using this substrate, we successfully amplified six sporadic Creutzfeldt-Jakob disease (sCJD) strains and the variant Creutzfeldt-Jakob disease (vCJD) strain from both human PrP transgenic (tg650) mouse brain homogenates and directly from patients\u2019 samples. In tg650-passaged prions, amplification reactions were initiated between 3 and 36 hours for sCJD prions and between 11 and 31 hours for vCJD prions, covering a 5- to 7-log dilution range depending on the strain. For patient brain homogenates, amplification reactions started between 0 and 27 hours for sCJDs and between 17 and 35 hours for vCJD, covering a 5- to 8-log dilution range depending on the strain. VV1 prions from a patient sample could only be amplified over a 2-log dilution range. Moreover, lag times of amplification reactions enabled reliable discrimination between vCJD and all tested sCJD subtypes. These findings represent a significant advance toward ante-mortem typing of human prion diseases.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>IMPORTANCE<\/jats:title>\n                    <jats:p>Creutzfeldt-Jakob disease (CJD) is a fatal neurodegenerative disorder within the prion disease family. Definitive diagnosis and strain typing currently require post-mortem analyses. In this study, we demonstrate that the human PrP E219K variant serves as an effective substrate for prion in vitro amplification using real-time quaking-induced conversion. This substrate enables (i) rapid and robust amplification of the most common human prion strains and (ii) clear and direct discrimination between variant CJD and all tested sporadic CJD subtypes, based on statistical analyses of lag times of the prion amplification reactions. These findings represent a significant step toward the development of ante-mortem tools for prion strain typing in affected patients.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.1128\/spectrum.00292-25","type":"journal-article","created":{"date-parts":[[2025,7,10]],"date-time":"2025-07-10T13:00:23Z","timestamp":1752152423000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":5,"title":["Human PrP E219K: a new and promising substrate for robust RT-QuIC amplification of human prions with potential for strain discrimination"],"prefix":"10.1128","volume":"13","author":[{"given":"A.","family":"Mar\u00edn-Moreno","sequence":"first","affiliation":[{"id":[{"id":"https:\/\/ror.org\/01jvz7e61","id-type":"ROR","asserted-by":"publisher"}],"name":"Universit\u00e9 Paris-Saclay, Institut National de Recherche pour l\u2019Agriculture, l\u2019Alimentation et l\u2019Environnement, Universit\u00e9 Versailles-Saint Quentin, Unit\u00e9 de Virologie et d\u2019Immunologie Mol\u00e9culaires","place":["Jouy-en-Josas, France"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"F.","family":"Reine","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/01jvz7e61","id-type":"ROR","asserted-by":"publisher"}],"name":"Universit\u00e9 Paris-Saclay, Institut National de Recherche pour l\u2019Agriculture, l\u2019Alimentation et l\u2019Environnement, Universit\u00e9 Versailles-Saint Quentin, Unit\u00e9 de Virologie et d\u2019Immunologie Mol\u00e9culaires","place":["Jouy-en-Josas, France"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"F.","family":"Jaffr\u00e9zic","sequence":"additional","affiliation":[{"name":"Universit\u00e9 Paris-Saclay, Institut National de Recherche pour l\u2019Agriculture, l\u2019Alimentation et l\u2019Environnement, AgroParisTech, Unit\u00e9 de G\u00e9n\u00e9tique Animale et Biologie Int\u00e9grative","place":["Jouy-en-Josas, France"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"L.","family":"Herzog","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/01jvz7e61","id-type":"ROR","asserted-by":"publisher"}],"name":"Universit\u00e9 Paris-Saclay, Institut National de Recherche pour l\u2019Agriculture, l\u2019Alimentation et l\u2019Environnement, Universit\u00e9 Versailles-Saint Quentin, Unit\u00e9 de Virologie et d\u2019Immunologie Mol\u00e9culaires","place":["Jouy-en-Josas, France"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"H.","family":"Rezaei","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/01jvz7e61","id-type":"ROR","asserted-by":"publisher"}],"name":"Universit\u00e9 Paris-Saclay, Institut National de Recherche pour l\u2019Agriculture, l\u2019Alimentation et l\u2019Environnement, Universit\u00e9 Versailles-Saint Quentin, Unit\u00e9 de Virologie et d\u2019Immunologie Mol\u00e9culaires","place":["Jouy-en-Josas, France"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"I.","family":"Quadrio","sequence":"additional","affiliation":[{"name":"Biochemistry and Molecular Biology Department, Neurodegenerative Pathologies, LBMMS, Hospices Civils de Lyon, H\u00f4pital neurologique","place":["Lyon, France"]},{"name":"BIORAN Team, Lyon Neurosciences Research Center","place":["Lyon, France"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"S.","family":"Ha\u00efk","sequence":"additional","affiliation":[{"name":"Sorbonne Universit\u00e9, Institut du Cerveau - 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