Journal
ANALYTICAL CHEMISTRY
Volume 86, Issue 17, Pages 8873-8880Publisher
AMER CHEMICAL SOC
DOI: 10.1021/ac502390c
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Funding
- National Natural Science Foundation of China [31171702]
- Project of National Science & Technology Pillar Plan [2012BAB19B09]
- National Key Project for Agro-product Quality & Safety Risk Assessment, PRC [GJFP2014006]
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A phage-displayed library of variable domain of heavy chain of the heavy chain antibody (VHH) or nanobody (Nb) was constructed after immunizing an alpaca with aflatoxin B1 (AFB(1)) conjugated with bovine serum albumin (AFB(1)-BSA). Two AFB(1)-specific nanobodies were selected. The obtained nanobodies were compared to an aflatoxin-specific monoclonal antibody B5 with respect to stability under organic solvents and high temperature. The two nanobodies could bind antigen specifically after exposure to temperatures as high as 95 degrees C. Besides, the nanobodies showed better or similar tolerance to organic solvents. A competitive ELISA with nanobody Nb26 was developed for the analysis of AFB(1), exhibiting an IC50 value of 0.754 ng/mL (2.4 mu M), linear range from 0.117 to 5.676 ng/mL. Due to the high tolerance to methanol, sample extracts were analyzed by nanobody-based ELISA without dilution. The recovery from spiked peanut, rice, corn and feedstuff ranged from 80 to 115%. In conclusion, the isolated nanobodies are excellent candidates for immunoassay application in aflatoxin determination.
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