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High-Throughput Automated Microwave-Enhanced Peptide Synthesis
Introduction
Modern peptide research is fast paced and often involves screening libraries of peptides to discover and develop functional peptides. Automated microwave-assisted SPPS has greatly reduced the time required to synthesize peptides, but synthesizing one peptide at a time requires user input between each sequence, if libraries need to be prepared. CEM’s microwave peptide synthesizers (Liberty LiteTM 2.0, Liberty BlueTM 2.0, and Liberty PRIMETM 2.0) can be paired with an HT transfer system, to allow the user to queue and automatically synthesize up to 24 peptides consecutively. Batches of individual resins are pre-loaded onto the HT for each peptide in the queue, and then automatically transferred to the Liberty synthesizer for peptide synthesis.
Automatic resin transfer removes the need for user input in between each queued peptide, maximizing productivity during the work-day and allows multiple peptides to run overnight. HT systems are designed to transfer 4, 12, or 24 batches of resin per queue, with ideal usage, based on the cycle times of the synthesizer. The fastest synthesizer, Liberty PRIME 2.0, can be equipped with HT4, HT12, or HT24, depending on the needs of the researcher. Liberty Blue 2.0 can be equipped with HT4, or HT12, while the base model Liberty Lite 2.0 can be equipped with an HT4. Furthermore, the Liberty PRIME 2.0 synthesizer features extra reagent and bottle capacity for running large batches with the HT24 setup, without refilling, along with the optional ability to be used in cGMP settings for peptide production.
To demonstrate, a published set of 20 neoantigen peptides with diverse sequences and an average length of 16 residues was synthesized with the Liberty PRIME 2.0 using an HT24. The total synthesis time for the set of peptides was 24 h 14 min. The peptides were synthesized on 0.1 mmol scale using CarboMAXTM and yielded crude peptides with purities that ranged from 47% – 90%, and an average purity of 69% (Table 1 on page 2 and Figure 1 on page 3). The total usage of DMF was 6056 mL and the total amount of waste generated was 6979 mL. These neo-antigens were further purified by elevated temperature chromatography to purities suitable for biological studies.