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Recombinant MHC II proteins are effective for the analysis of peptide\u2010binding interaction, detection of antigen\u2010specific T\u2010cells and clinical testing for donor\u2010specific anti\u2010human leukocyte antigen (HLA) antibody for organ transplantation. However,\n                    <jats:styled-content style=\"fixed-case\">\n                      <jats:italic>Escherichia coli<\/jats:italic>\n                    <\/jats:styled-content>\n                    \u2010based production is limited due to difficulty in obtaining properly folded protein complexes. In this study, we utilised the expression machinery of\n                    <jats:italic>Nicotiana benthamiana<\/jats:italic>\n                    via the Tsukuba System to produce the soluble extracellular domains of MHC II proteins. Thirteen different MHC II proteins of the three isotypes (HLA\u2010DR, DQ and DP) conjugated with peptides (pMHC II) were successfully expressed. The HLA\u2010DRA*01:01\/DRB1*01:01 covalently linked to a class II\u2010associated invariant chain peptide (CLIP) or influenza hemagglutinin (HA, 306\u2013318) was further analysed as a model. The resultant proteins were detectable in soluble fractions and purified using metal affinity purification and ion exchange chromatography, yielding 4.37 and 2.81\u2009\u03bcg\/gFW, respectively. pMHC II were also biotinylated in\u00a0vitro using commercial BirA, enabling future construction of tetramers or multimers. The purified proteins were confirmed through enzyme\u2010linked immunosorbent assay (ELISA) using anti\u2010HLA antibodies. Furthermore, Surface plasmon resonance biosensor (Biacore) verified glycosylation\u2010dependent recognition of MHC II\u2010HA complex by cognate T\u2010cell receptor (HA1.7 TCR), resulting in an equilibrium dissociation constant of K\n                    <jats:sub>D<\/jats:sub>\n                    \u2009=\u20093.62\u2009\u00b1\u20091.59\u2009\u03bcM. Overall, this plant\u2010based system offers a promising scalable and economical alternative for large\u2010scale production of functional MHC II proteins for basic and clinical applications.\n                  <\/jats:p>","DOI":"10.1111\/pbi.70705","type":"journal-article","created":{"date-parts":[[2026,7,1]],"date-time":"2026-07-01T12:10:45Z","timestamp":1782907845000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Plant\u2010Based Production of Human Major Histocompatibility Complex Class\n                    <scp>II<\/scp>\n                    Molecules"],"prefix":"10.1111","author":[{"given":"Abdelaziz","family":"Ramadan","sequence":"first","affiliation":[{"name":"Degree Programs in Life and Earth Sciences University of Tsukuba  Tsukuba Japan"},{"name":"Tsukuba\u2010Plant Innovation Research Center University of Tsukuba  Tsukuba Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hiroko","family":"Miyadera","sequence":"additional","affiliation":[{"name":"Department of Medical Genetics, Institute of Medicine University of Tsukuba  Ibaraki Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kaho","family":"Oka","sequence":"additional","affiliation":[{"name":"Degree Programs in Life and Earth Sciences University of Tsukuba  Tsukuba Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kohei","family":"Ishikawa","sequence":"additional","affiliation":[{"name":"Tokyo Power Technology Ltd.  Chiba Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Toshiya","family":"Kobayashi","sequence":"additional","affiliation":[{"name":"Tokyo Power Technology Ltd.  Chiba Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Misaki","family":"Kobayashi","sequence":"additional","affiliation":[{"name":"Degree Programs in Life and Earth Sciences University of Tsukuba  Tsukuba Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3133-157X","authenticated-orcid":false,"given":"Mitsunori","family":"Shiroishi","sequence":"additional","affiliation":[{"name":"Department of Biological Science and Technology Tokyo University of Science  Tokyo Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Emiko","family":"Noguchi","sequence":"additional","affiliation":[{"name":"Department of Medical Genetics, Institute of Medicine University of Tsukuba  Ibaraki Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1262-2176","authenticated-orcid":false,"given":"Kenji","family":"Miura","sequence":"additional","affiliation":[{"name":"Degree Programs in Life and Earth Sciences University of Tsukuba  Tsukuba Japan"},{"name":"Tsukuba\u2010Plant Innovation Research Center University of Tsukuba  Tsukuba Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2026,7,1]]},"reference":[{"key":"e_1_2_9_2_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.coi.2021.04.009"},{"key":"e_1_2_9_3_1","doi-asserted-by":"publisher","DOI":"10.1038\/381616a0"},{"key":"e_1_2_9_4_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.ymeth.2012.03.025"},{"key":"e_1_2_9_5_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0022-1759(02)00200-4"},{"key":"e_1_2_9_6_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.tplants.2020.04.009"},{"key":"e_1_2_9_7_1","doi-asserted-by":"publisher","DOI":"10.1002\/biot.201200032"},{"key":"e_1_2_9_8_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.91.24.11408"},{"key":"e_1_2_9_9_1","doi-asserted-by":"publisher","DOI":"10.4049\/jimmunol.178.9.5727"},{"key":"e_1_2_9_10_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41587\u2010021\u201000893\u20109"},{"key":"e_1_2_9_11_1","doi-asserted-by":"publisher","DOI":"10.1007\/978\u20101\u20104939\u20102272\u20107_12"},{"key":"e_1_2_9_12_1","doi-asserted-by":"publisher","DOI":"10.1159\/000237390"},{"key":"e_1_2_9_13_1","first-page":"310","article-title":"Prediction 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