A one-step immunoassay based on switching peptides for diagnosis of porcine epidemic diarrhea virus (PEDV) using screened Fv-antibodies

• Published in: Journal of materials chemistry. B, Materials for biology and medicine Materials for biology and medicine, 2024-04, Vol.12 (15), p.3751-3763
• Main Authors: Kim, Tae-Hun, Park, Jae-Yeon, Jung, Jaeyong, Sung, Jeong Soo, Kwon, Soonil, Bae, Hyung Eun, Shin, Hyun-Jin, Kang, Min-Jung, Jose, Joachim, Pyun, Jae-Chul
• Format: Article
• Language: English
• Subjects: ACE2; Affinity; Amino acid sequences; Amino acids; Aminopeptidase; Angiotensin; Angiotensin-converting enzyme 2; Antibodies; Antibody libraries; Antigens; Binding; Constants; Diarrhea; Epidemics; Fluorescence; Immunoassay; Immunoassays; Peptides; Peptidyl-dipeptidase A; Proteins; Receptors; Spike protein; Switching; Transmissible gastroenteritis; Viruses
• ISSN: 2050-750X; 2050-7518
• DOI: 10.1039/d4tb00066h

In this study, a one-step immunoassay for porcine epidemic diarrhea virus (PEDV) based on Fv-antibodies and switching peptides was developed, and the assay results of PEDV were obtained by just mixing samples without any further reaction or washing steps. The Fv-antibodies with binding affinity to the spike protein of PEDV were screened from the Fv-antibody library using the receptor-binding domain (RBD) of the spike protein as a screening probe. Screened Fv-antibodies with binding affinities to the RBD antigen were expressed, and the binding constants ( K D ) were calculated to be 83-142 nM. The one-step immunoassay for the detection of PEDV was configured as a displacement immunoassay using a fluorescence-labeled switching peptide. The one-step immunoassay based on switching peptides was performed using PEDV, and the limit of detection (LOD) values for PEDV detection were estimated to be Ct = 39.7-36.4. Compared with the LOD value for a conventional lateral flow immunoassay (Ct = 33.0), the one-step immunoassay showed a remarkably improved LOD for the detection of PEDV. Finally, the interaction between the screened Fv-antibodies and the PEDV RBD was investigated using docking simulations and compared with the amino acid sequences of the receptors on host cells, such as aminopeptidase N (APN) and angiotensin-converting enzyme-2 (ACE-2). In this study, a one-step immunoassay for porcine epidemic diarrhea virus (PEDV) based on Fv-antibodies and switching peptides was developed, and the assay results of PEDV were obtained by just mixing samples without any further reaction or washing steps.