Efficient Strategy\nto Discover DNA Aptamers Against\nLow Abundance Cell Surface Proteins in Scarce Samples

Molecular recognition probes targeting cell surface proteins\nsuch\nas aptamers play crucial roles in precise diagnostics and therapy.\nHowever, the selection of aptamers against low-abundance proteins\nin situ on the cell surface, especially in scarce samples, remains\nan unmet challenge. In this study, we present a single-round, single-cell\naptamer selection method by employing a digital DNA sequencing strategy,\ntermed DiDS selection, to address this dilemma. This approach incorporates\na molecular identification card for each DNA template, thereby mitigating\nbiases introduced by multiple PCR amplifications and ensuring the\naccurate identification of aptamer candidates. Through DiDS selection,\nwe successfully obtained a series of high-quality aptamers against\ncell lines, clinical specimens, and neurons. Subsequent analyses for\ntarget identification revealed that aptamers derived from DiDS selection\nexhibit recognition capabilities for proteins with varying abundance\nlevels. In contrast, multiple rounds of selection resulted in the\nenrichment of only one aptamer targeting a high-abundance target.\nMoreover, the comprehensive profiling of cell surfaces at the single-cell\nlevel, utilizing an enriched aptamer pool, revealed unique molecular\npatterns for each cell line. This streamlined approach holds promise\nfor the rapid generation of specific recognition molecules targeting\ncell surface proteins across a broad range of expression levels and\nexpands its applications in cell profiling, specific probe identification,\nbiomarker discovery, etc.