Screening specific aptamer pairs towards bovine lactoferrin for its sandwich application
Lactoferrin (Lf) is closely associated with the growth of infants and children, and its recognition is imperative due to its rising application as a prevalent nutritional fortification in infant formula. In this study, an ssDNA aptamer was screened out that enabled sandwich recognition of Lf by forming an aptamer pair with another available aptamer. During the process, the reported aptamer (SeqT1.1) was selected as the substrateto be linked onto pre-blocked microplates for capturing Lf, followed by the addition of the ssDNA library, resulting in the formation of the sandwich complex of the “SeqT1.1/Lf/ssDNA library” that was subsequently collected for screening. After seven rounds, we discovered a new ssDNA aptamer (Seq6T) through a comprehensive analysis involving sequence frequency, secondary structures, homology, and sequence truncation, which helped address the binding interference between aptamer pairs with each other’s binding. The Seq6T demonstrated good specificity and low-nanomolar affinity by ELONA (K D , 12.59 nM) and CE-LIF (K D , 22.44 nM) assays, which illustrated the potential of applying that in both homogeneous and non-homogeneous environments. The sandwich recognition of “SeqT1.1/Lf/Seq6T” was further verified by molecular docking and ELONA, with a good detection limit of 18.35 nM. The aptamer pairs provided a considerable basis for accelerating the sandwich recognition applications towards Lf.
危险品化学品经营许可证（带存储）