Standard
solid-phase N-αFmoc chemistry was used to synthesize meditope
derivatives on the CS136XT peptide synthesizer (C S BIO). After cleavage
of the peptides from resin using reagent K (TFA/water/phenol/thioanisole/EDT
= 82.5:5:5:5:2.5), crude peptides were collected by precipitation
from cold ether. For disulfide-linked meditopes, a further oxidation
using either 20% DMSO in ammonium acetate buffer (pH 6) or iodine
was performed. All peptides were purified using a reverse-phase HPLC
(Agilent 1200 system with Agilent prep-C18 column, 21.2 × 150
mm, 5 μm) with a water (0.1% TFA)/acetonitrile (0.1% TFA) solvent
system. All peptides were characterized by mass spectrometry. Alexa
Fluor 647 labeled peptides were synthesized from Alexa Fluor 647-NHS
(Thermo Fisher Scientific, Waltham, MA). Reverse-phase HPLC purification
provided the purified M-A647: Ac-CQFDXSTRRLRCGGSK-A647 (X = diphenylalanine).34 (link)
solid-phase N-αFmoc chemistry was used to synthesize meditope
derivatives on the CS136XT peptide synthesizer (C S BIO). After cleavage
of the peptides from resin using reagent K (TFA/water/phenol/thioanisole/EDT
= 82.5:5:5:5:2.5), crude peptides were collected by precipitation
from cold ether. For disulfide-linked meditopes, a further oxidation
using either 20% DMSO in ammonium acetate buffer (pH 6) or iodine
was performed. All peptides were purified using a reverse-phase HPLC
(Agilent 1200 system with Agilent prep-C18 column, 21.2 × 150
mm, 5 μm) with a water (0.1% TFA)/acetonitrile (0.1% TFA) solvent
system. All peptides were characterized by mass spectrometry. Alexa
Fluor 647 labeled peptides were synthesized from Alexa Fluor 647-NHS
(Thermo Fisher Scientific, Waltham, MA). Reverse-phase HPLC purification
provided the purified M-A647: Ac-CQFDXSTRRLRCGGSK-A647 (X = diphenylalanine).34 (link)
