VHH phage-based competitive real-time immuno-polymerase chain reaction for ultrasensitive detection of ochratoxin A in cereal

Details

• Personal Author:
  Fu JH ; Gee SJ ; Hammock BD ; He ZY ; Li YP ; Liu X ; Qiu YL ; Tu Z ; Xiong YH ; Xu Y
• Description:
  Phage display-mediated immuno-polymerase chain reaction (PD-IPCR) is an ultrasensitive detection technology that combines the advantages of immuno-PCR and phage display. The phage particle, which displayed antibody fragments including single-chain fragment variable (scFv), variable domain of heavy-chain antibodies (VHH), and antigen-binding fragment (Fab) on the surface can be directly used in IPCR, supplying both the detection antibody and deoxyribonucleic acid (DNA) template. In this work, we used ochratoxin A (OTA) as a model system to study the capacity of PD-IPCR in the detection of toxic small molecular weight compounds, especially mycotoxins. An alpaca-derived VHH library was constructed and subjected to four cycles of panning. In total, 16 clones with four unique sequences were selected by competitive binding with OTA. The clone VHH-28 resulted in the lowest 50% inhibitory concentration of 0.31 ng/mL in the phage enzyme-linked immunosorbent assay (ELISA) and was selected to develop the VHH phage-based real-time immuno-PCR (RT-IPCR). The detection limit of the VHH phage-based RT-IPCR was 3.7 pg/L, with a linear range of 0.01-1000 pg/mL. This method was compared with conventional ELISA, and validation results indicated the reliability of VHH phage-based RT-IPCR in the detection of OTA in cereal samples. This study provides a new idea for the ultrasensitive detection of mycotoxins and other toxic small molecular weight compounds. [Description provided by NIOSH]
• Subjects:
  Carcinogens Energy Metabolism Food Contamination Fungi Hazardous Substances Kidney Diseases Nervous System Occupational Health Safety Teratogenesis Urogenital Diseases Urogenital System

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• Keywords:
  Contaminated-food Food Food-contaminants Foodstuff Metabolites Nephrotoxicity Nephrotoxins Neurotoxic-effects Teratogens Toxins
• ISSN:
  0003-2700
• Document Type:
  Text
• Funding:
  Cooperative Agreement
• Genre:
  Journal Article
• Place as Subject:
  California ; OSHA Region 9
• CIO:
  National Institute for Occupational Safety and Health (NIOSH)
• Topic:
  Workplace Safety & Health
• Location:
  North America
• Volume:
  86
• Issue:
  15
• NIOSHTIC Number:
  nn:20046657
• Citation:
  Anal Chem 2014 Aug; 86(15):7471-7477
• Contact Point Address:
  Yang Xu, Sino-Germany Joint Research Institute, Nanchang University, 235 Nanjing East Road, Nanchang 330047, People's Republic of China
• Email:
  xuyang@ncu.edu.cn
• Federal Fiscal Year:
  2014
• NORA Priority Area:
  Agriculture, Forestry and Fishing
• Performing Organization:
  University of California - Davis
• Peer Reviewed:
  True
• Start Date:
  20010930
• Source Full Name:
  Analytical Chemistry
• End Date:
  20270929
• Collection(s):
  National Institute for Occupational Safety and Health
• Main Document Checksum:
  urn:sha-512:eb5c48a8aaf7b1509f5f46e0fff14ca00bfb732e6895502eb56216739ee7ae076081227cba2af0648ec843b9b789d0d282017af4a09e808a8d0e1078257fa2e1
• Download URL:
  https://stacks.cdc.gov/view/cdc/202016/cdc_202016_DS1.pdf
• File Type:
  [PDF - 2.26 MB ]