An ultrasensitive bioluminescent enzyme immunoassay based on nanobody/nanoluciferase heptamer fusion for the detection of Tetrabromobisphenol A in sediment
Li, Z.; Wang, Y.; Vasylieva, N.; Wan, D.; Yin, Z.; Dong, J.; Hammock, B.D. (2020). An ultrasensitive bioluminescent enzyme immunoassay based on nanobody/nanoluciferase heptamer fusion for the detection of Tetrabromobisphenol A in sediment. Anal. Chem. 92(14): 10083-10090. https://dx.doi.org/10.1021/acs.analchem.0c01908
In: Analytical chemistry. American Chemical Society: Washington. ISSN 0003-2700; e-ISSN 1520-6882, more
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| Authors | | Top |
- Li, Z.
- Wang, Y.
- Vasylieva, N.
- Wan, D.
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- Yin, Z.
- Dong, J.
- Hammock, B.D.
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| Abstract |
Tetrabromobisphenol A (TBBPA) is a flame retardant and has become a widely concerning environmental pollutant. An ultrasensitive nanobody-based immunoassay was developed to monitor the exposure of TBBPA in sediment. First, the anti-TBBPA nanobody was fused with nanoluciferase, and then a one-step bioluminescent enzyme immunoassay (BLEIA) was developed with high sensitivity for TBBPA, with a maximum half inhibition concentration (IC50) at 187 pg/mL. Although approximately 10-fold higher sensitivity can be achieved by this developed BLEIA than by the classical two-step ELISA (IC50 at 1778 pg/mL), it is still a challenge to detect trace TBBPA in sediment samples reliably due to the relatively high matrix effect. To further improve the performance of this one-step BLEIA, a C4b-binding protein (C4BP) was inserted as a self-assembling linker between the nanobody and nanoluciferase. Therefore, a heptamer fusion containing seven binders and seven tracers was generated. This reagent improved the binding capacity and signal amplification. The one-step heptamer plus BLEIA based on this immune-reagent shows an additional 7-fold improvement of sensitivity, with the IC50 of 28.9 pg/mL and the limit of detection as low as 2.5 pg/mL. The proposed assay was further applied to determine the trace TBBPA in sediment, and the recovery was within 92–103%. Taking advantage of this heptamer fusion, one-step BLEIA can serve as a powerful tool for fast detection of trace TBBPA in the sediment samples. |
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