High Throughput pMHC-I Multimer Library Production Using Chaperone-Mediated Peptide Exchange
| العنوان: | High Throughput pMHC-I Multimer Library Production Using Chaperone-Mediated Peptide Exchange |
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| المؤلفون: | Sarah Overall, Jugmohit S Toor, Mark Yarmarkovich, Stephanie Hao, Son Nguyen, Alberto Sada-Japp, Michael R Betts, John M Maris, Peter Smibert, Nikolaos G Sgourakis |
| المصدر: | The Journal of Immunology. 202:130.4-130.4 |
| بيانات النشر: | The American Association of Immunologists, 2019. |
| سنة النشر: | 2019 |
| مصطلحات موضوعية: | Immunology, Immunology and Allergy |
| الوصف: | The development of pMHC-multimer technologies has been indispensable for the characterisation of T cell responses to individual viral and tumor peptide antigens. Given the large variety of T cell receptors expressed in polyclonal repertoires, peptide exchange technologies are becoming increasingly important in providing a high throughput means of generating pMHC-multimer libraries, containing a wide range of antigens to probe such repertoires. Here we present a simple method for the isolation of empty MHC-I molecules which can be readily used for high throughput multimer library preparation. We utilise the chaperone TAPBPR, which binds MHC-I molecules and stabilises them in a peptide-receptive conformation, to isolate empty MHC-I molecules through the combination of placeholder peptides and small molecule competitors. We demonstrate that empty H-2Dd:TAPBPR and HLA-A*02:01:TAPBPR complexes can be stored long term and subsequently loaded with peptides of choice in a high throughput fashion to afford pMHC-multimer libraries. The resulting pMHC-multimers show equivalent flow cytometric properties to multimers made from MHC-I molecules refolded in vitro with synthetic peptides and are significantly improved relative to multimers prepared using peptide exchange of conditional ligands. Using this simple system to generate a library of oligonucleotide-labeled pMHC-multimers, combined with our recently described multi-modal cellular indexing technology (ECCITE-seq), we can identify distinct T cell receptor sequences present in polyclonal repertoires, together with their antigen specificities. |
| تدمد: | 1550-6606 0022-1767 |
| الوصول الحر: | https://explore.openaire.eu/search/publication?articleId=doi_________::b6136dbb273d65bb1735fe9778d9a359Test https://doi.org/10.4049/jimmunol.202.supp.130.4Test |
| رقم الانضمام: | edsair.doi...........b6136dbb273d65bb1735fe9778d9a359 |
| قاعدة البيانات: | OpenAIRE |
| تدمد: | 15506606 00221767 |
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