|
| Status |
Public on Mar 12, 2020 |
| Title |
High Throughput pMHC-I Tetramer Library Production Using Chaperone Mediated Peptide Exchange |
| Organisms |
Homo sapiens; synthetic construct |
| Experiment type |
Expression profiling by high throughput sequencing
|
| Summary |
Peptide exchange technologies are essential for the generation of pMHC-multimer libraries for probing diverse, polyclonal TCR repertoires in various settings. Here we report, using the molecular chaperone TAPBPR, a robust method for the capture of stable, empty MHC-I molecules, which can be readily tetramerized and loaded with peptides of choice in a high-throughput manner. Alternatively, catalytic amounts of TAPBPR can be used to exchange ‘goldilocks’ peptides with high-affinity peptides of interest in an overnight reaction. Using the same system, we describe high-throughput assays to validate binding of multiple candidate peptides on the empty MHCI groove. Combined with tetramer-barcoding via a multi-modal cellular indexing technology, ECCITE-seq, our approach allows a combined analysis of TCR repertoires and other T-cell transcription profiles together with their cognate pMHC-I specificities in a single experiment.
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| |
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| Overall design |
spleen, jurkat, and neuroblastoma samples
|
| |
|
| Contributor(s) |
Hao S, Smibert P |
| Citation(s) |
32312993 |
| |
| Submission date |
Mar 11, 2020 |
| Last update date |
Apr 27, 2020 |
| Contact name |
Peter Smibert |
| E-mail(s) |
psmibert@nygenome.org
|
| Organization name |
New York Genome Center
|
| Street address |
101 Avenue of the Americas
|
| City |
New York |
| State/province |
NY |
| ZIP/Postal code |
10013 |
| Country |
USA |
| |
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| Platforms (2) |
| GPL15520 |
Illumina MiSeq (Homo sapiens) |
| GPL17769 |
Illumina MiSeq (synthetic construct) |
|
| Samples (5)
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|
| Relations |
| BioProject |
PRJNA612056 |
| SRA |
SRP252454 |
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