Synthetic Yellow Fever Peptide Epitope

The yellow fever specific peptide epitope K9F (KTWGKNLVF) and two synthetic variants, K9A (KTWGKNLVA) and K9V (KTWGKNLVV), were synthesized by standard 9-fluorenylmethyloxycarbonyl (FMOC) chemistry and purified by reversed-phase high-performance liquid chromatography (purity at least 80%, usually >95%) (Schafer-N, Copenhagen, Denmark). Tetramers were produced as previously described [21 (link)] with the following modification: abacavir (ABC; 200 μg/ml) was added to the folding reaction for the production of the variant tetramers, K9A-ABC-B*57:01 and K9V-ABC-B*57:01. Successfully folding of MHC-I monomers with these peptides was strictly abacavir dependent. To tetramerize the complexes Streptavidin-R-Phycoerythin (PE), Streptavidin-Allophycocyanin (APC), or Streptavidin-Brilliant Violet 421 (BV421) (Biolegend, San Diego, USA) was sequentially added over 60 min at a molar ratio of Streptavidin to peptide-MHC-I monomer of 1:4.

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Lucas A., Lucas M., Strhyn A., Keane N.M., McKinnon E., Pavlos R., Moran E.M., Meyer-Pannwitt V., Gaudieri S., D’Orsogna L., Kalams S., Ostrov D.A., Buus S., Peters B., Mallal S, & Phillips E. (2015). Abacavir-Reactive Memory T Cells Are Present in Drug Naïve Individuals. PLoS ONE, 10(2), e0117160.

Publication 2015

Streptavidin-Allophycocyanin (APC),

Abacavir Allophycocyanin Complexes r Epitope High performance liquid chromatography Mhc i peptide Molar Peptides Streptavidin Tetramers Violet Yellow fever

Corresponding Organization : La Jolla Institute For Allergy & Immunology

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Variable analysis

independent variables

Peptide epitopes: K9F, K9A, and K9V

dependent variables

Successful folding of MHC-I monomers with the peptides

control variables

Abacavir (ABC; 200 μg/ml) was added to the folding reaction for the production of the variant tetramers, K9A-ABC-B*57:01 and K9V-ABC-B*57:01

controls

Positive control: Not explicitly mentioned.
Negative control: Not explicitly mentioned.

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