Rapid single-step upconversion-linked immunosorbent assay for diclofenac
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The non-steroidal anti-inflammatory drug and analgesic diclofenac is a common micropollutant in water. A direct competitive upconversion-linked immunosorbent assay (ULISA) for diclofenac has been developed that is based on a nanoparticulate tracer consisting of a NaYF4:Yb,Er core (diameter of ~90 nm, excitation and emission wavelenghts 980 and 535 nm, respectively) enclosed by a carboxylated silica shell and finally coated with diclofenac-conjugated bovine γ-globulin. The proteinaceous coating prevents non-specific adsorption of the tracer to the microtiter plate and provides a structurally flexible linker for surface-exposed diclofenac to warrant efficient competition with free (analyte) diclofenac in real water samples. The tracer was purified by gel electrophoresis and lyophilized. Both processes have no adverse effects on the immunoassay. All assay components can be stored in a dry state without a cooling chain, and can be reactivated on demand. Hence, this ULISA is well suited for environmental monitoring in low-resource settings. The ULISA has a similar limit of detection (20 pg mL−1; equivalent to 70 pM) as the conventional ELISA, but the time for analysis is reduced to 70 min because no enzymatic amplification steps are involved.
KeywordsBioconjugation Electrophoretic purification Immunoassay Luminescence Pharmaceutical micropollutant Photon-upconversion
We acknowledge financial support from the COST Action CM1403 “The European Upconversion Network: From the Design of Photon-Upconverting Nanomaterials to Biomedical Applications”. H.H.G. acknowledges funding from the German Research Foundation for a Heisenberg Fellowship (DFG, Grant GO 1968/5-1 and GO 1968/6-1). Further funding was provided by the Czech Ministry of Education, Youth and Sports (COST CZ project LD15023), ANR-DFG program (project NArBioS, Grant no. ANR-11-INTB-1013) and the Grant Agency of the Czech Republic (P20612G014 GACR). CIISB research infrastructure project LM2015043 funded by MEYS CR is gratefully acknowledged for the financial support of the measurements at the CF Cryo-electron Microscopy and Tomography.
Compliance with ethical standards
The author(s) declare that they have no competing interests.
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