An immunized mouse phage display scFv library with a capacity of 3.34 × 109 CFU/mL was constructed and used for screening of recombinant anti-ciprofloxacin single-chain antibody for the detection of ciprofloxacin (CIP) in animal-derived food. After four rounds of bio-panning, 25 positives were isolated and identified successfully. The highest positive scFv-22 was expressed in E. coli BL21. Then, its recognition mechanisms were studied using the molecular docking method. The result showed the amino acid residue Val160 was the key residue for the binding of scFv to CIP. Based on the results of virtual mutation, the scFv antibody was evolved by directional mutagenesis of contact amino acid residue Val160 to Ser. After the expression and purification, an indirect competitive enzyme-linked immunosorbent assay (IC-ELISA) based on the parental and mutant scFv was established for CIP, respectively. The IC50 value of the assay established with the ScFv mutant was 1.58 ng/mL, while the parental scFv was 26.23 ng/mL; this result showed highly increased affinity, with up to 16.6-fold improved sensitivity. The mean recovery for CIP ranged from 73.80% to 123.35%, with 10.46% relative standard deviation between the intra-assay and the inter-assay. The RSD values ranged between 1.49% and 9.81%. The results indicate that we obtained a highly sensitive anti-CIP scFv by the phage library construction and directional evolution, and the scFv-based IC-ELISA is suitable for the detection of CIP residue in animal-derived edible tissues.
DPL ( http://www.peptide-ligand.cn/ ) is a comprehensive database of peptide ligand (DPL). DPL1.0 holds 1044 peptide ligand entries and provides references for the study of the polypeptide platform. The data were collected from PubMed-NCBI, PDB, APD3, CAMPR3, etc. The lengths of the base sequences are varied from 3 to78. DPL database has 923 linear peptides and 88 cyclic peptides. The functions of peptides collected by DPL are very wide. It includes 540 entries of antiviral peptides (including SARS-CoV-2), 55 entries of signal peptides, 48 entries of protease inhibitors, 45 entries of anti-hypertension, 37 entries of anticancer peptides, etc. There are 270 different kinds of peptide targets. All peptides in DPL have clear binding targets. Most of the peptides and receptors have 3D structures experimentally verified or predicted by CYCLOPS, I-TASSER and SWISS-MODEL. With the rapid development of the COVID-2019 epidemic, this database also collects the research progress of peptides against coronavirus. In conclusion, DPL is a unique resource, which allows users easily to explore the targets, different structures as well as properties of peptides.
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