As the field of traditional DNA-Encoded Library (DEL) chemistry reaches maturity, expectations for library quality have become increasingly exacting. Beyond conventional optimizations focused on purification protocols and reaction yields, a growing number of research groups have pioneered solid-phase synthesis strategies to enhance peptide library purity. Here, we present and discuss the key insights gleaned from three recent publications on solid-phase DEL derivatives.

Shiyu Chen et al. pioneered a solid-phase purification strategy for DNA-encoded peptide libraries (PDELs) by engineering a modified Fmoc (mFmoc) protecting group equipped with a terminal azido1. This design enables the specific immobilization of desired peptide intermediates onto alkyne-functionalized controlled pore glass (CPG) beads via copper-free click chemistry following each coupling step (Fig. 1). After rigorous washing to eliminate unreacted building blocks and truncated byproducts, the pure products are released through standard Fmoc deprotection. This "capture-and-release" cycle successfully facilitated the construction of the longest reported five-round PDEL with purity exceeding 95%, effectively breaking the conventional four-round synthesis barrier. (Solid DEL-1)

Fig. 1: Iterative cycles of generating a purified DNA-encoded peptide library with mFmoc-protected amino acids. The desired DNA-encoded peptide is isolated after immobilization and purification (Solid DEL-1).

The group of Jörg Scheuermann developed another dual-linker solid-phase synthesis strategy on magnetic beads to achieve "self-purifying" release of DELs (Fig. 2)2. This solid-phase platform not only facilitates the synthesis of high-purity five-cycle desired peptide compounds but also significantly expands the compatible reaction scope to include water-free conditions, enabling transformations previously inaccessible in traditional aqueous DEL synthesis, such as the SnAP cyclization reaction and acid-mediated Boc-deprotection. However, this synthetic strategy is quite tedious. (Solid DEL-2)

Fig. 2: Synthetic strategy used for "self-purifying" release of DEL (Solid DEL-2).

Brian M. Paegel has pioneered an alternative solid-phase DNA-encoded library (DEL) synthesis strategy that integrates the "one-bead-one-compound" (OBOC) approach3. In this method, library members are constructed on solid-phase microbeads and linked to DNA tags via a photocleavable linker. This design facilitates the physical isolation and light-triggered release of desired peptide compounds, thereby enabling a broader spectrum of screening modalities, including activity-based assays and cellular phenotypic screening. However, this approach is inherently limited to a library size of 104–106 members, as it relies on the individual screening of compounds on discrete physical beads (Fig. 3). (Solid DEL-3)

Fig. 3: Synthetic strategy used for one-bead-one-compound of DEL (Solid DEL-3).

Collectively, these three solid-phase DEL design paradigms offer new insights into the future of peptide DEL library construction. They suggest that we can strategically leverage emerging technologies to fundamentally enhance peptide DEL library quality. Aligning with this evolving paradigm, HitGen is also exploring the introduction of novel solid-phase methodologies to elevate the quality of our peptide libraries. We anticipate that, in the near future, these innovations will be successfully translated into practice, pointing a new direction for the next generation of DEL synthesis.

Reference:

1. He Q, Wang Y, Tang X, et al. Enhanced screening via a pure DNA-encoded peptide library enabled by an Fmoc modification. Proc Natl Acad Sci U S A. 2026;123(8):e2524999123. doi:10.1073/pnas.2524999123

2. Keller M, Petrov D, Gloger A, et al. Highly pure DNA-encoded chemical libraries by dual-linker solid-phase synthesis. Science. 2024;384(6701):1259-1265. doi:10.1126/science.adn3412

3. Dixit A, Paegel BM. Solid-phase DNA-encoded library synthesis: a master builder's instructions. Nat Protoc. 2026;21(2):542-581. doi:10.1038/s41596-025-01190-4