TY - JOUR
T1 - Application of CdTe/CdS/ZnS quantum dot in immunoassay for aflatoxin B1 and molecular modeling of antibody recognition
AU - Zhang, Fuyuan
AU - Liu, Bing
AU - Zhang, Yan
AU - Wang, Junping
AU - Lu, Yang
AU - Deng, Juan
AU - Wang, Shuo
PY - 2019/1/24
Y1 - 2019/1/24
N2 - In order to develop a sensitive immunoassay for Aflatoxin B1 (AFB1) monitoring, a hybridoma secreting anti-AFB1 monoclonal antibody with high binding affinity was screened. A new type of CdTe/CdS/ZnS quantum dot was synthesized and conjugated with an artificial antigen for use as a fluorescent probe in a simple one-step fluorescence immunoassay (FLISA). The developed FLISA allowed a sensitive determination of AFB1 in cereal samples in a wide linear range, from 0.08 to 1.97 ng mL −1 , with a detection limit of 0.01 ng mL −1 in cereal samples. The corresponding immunoglobulin genes of the Fab fragment were cloned and sequenced, and expression of Fab was successfully verified in HEK293 cells, with a K D value of 1.09 × 10 −7 mol L −1 for AFB1. To investigate the interactions between the antibody and AFB1, molecular docking, molecular dynamic simulation, and quantum-chemical computation were performed on AFB1 and a homology model of the Fab fragment. Our results showed that residues Ser32, Trp93, and Trp98 played the most important roles in the binding through hydrogen bonds formation, Pi-Pi stacked/Pi-alkyl interactions, and van der Waals interactions. In addition, the electrostatic potential study of AFB1 demonstrated that electrostatic interactions also played an important role in the recognition process. Results from theoretical studies provide guidance for hapten design and antibody improvement through genetic engineering.
AB - In order to develop a sensitive immunoassay for Aflatoxin B1 (AFB1) monitoring, a hybridoma secreting anti-AFB1 monoclonal antibody with high binding affinity was screened. A new type of CdTe/CdS/ZnS quantum dot was synthesized and conjugated with an artificial antigen for use as a fluorescent probe in a simple one-step fluorescence immunoassay (FLISA). The developed FLISA allowed a sensitive determination of AFB1 in cereal samples in a wide linear range, from 0.08 to 1.97 ng mL −1 , with a detection limit of 0.01 ng mL −1 in cereal samples. The corresponding immunoglobulin genes of the Fab fragment were cloned and sequenced, and expression of Fab was successfully verified in HEK293 cells, with a K D value of 1.09 × 10 −7 mol L −1 for AFB1. To investigate the interactions between the antibody and AFB1, molecular docking, molecular dynamic simulation, and quantum-chemical computation were performed on AFB1 and a homology model of the Fab fragment. Our results showed that residues Ser32, Trp93, and Trp98 played the most important roles in the binding through hydrogen bonds formation, Pi-Pi stacked/Pi-alkyl interactions, and van der Waals interactions. In addition, the electrostatic potential study of AFB1 demonstrated that electrostatic interactions also played an important role in the recognition process. Results from theoretical studies provide guidance for hapten design and antibody improvement through genetic engineering.
KW - Aflatoxin B1
KW - Antibody recognition
KW - Fluoroimmunoassay
KW - Molecular modeling
KW - Quantum dot
KW - Recombinant antibody
UR - http://www.scopus.com/inward/record.url?scp=85054197636&partnerID=8YFLogxK
U2 - 10.1016/j.aca.2018.09.058
DO - 10.1016/j.aca.2018.09.058
M3 - Article
C2 - 30567644
AN - SCOPUS:85054197636
VL - 1047
SP - 139
EP - 149
JO - Analytica Chimica Acta
JF - Analytica Chimica Acta
SN - 0003-2670
ER -