Nanobody multimerization strategy to enhance the sensitivity of competitive ELISA for detection of ochratoxin A in coffee samples

Bao, Kunlu; Liu, Xing; Xu, Qi; Su, Bencao; Liu, Zilong; Cao, Hongmei; Chen, Qi

doi:10.1016/j.foodcont.2021.108167

Cited by 36 publications

(9 citation statements)

References 37 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The Nb28-Nluc fusion protein was produced by autoinduction expression, according to our previous work [ 34 ]. First, the recombinant expression plasmid pET25b-Nb28-Nluc was transformed into chemically competent cells ( E. coli Rosetta) by heat treatment at 42 °C for 90 s. Then, the transformed E. coli Rosetta cells were cultured in 300 mL of autoinduction medium at 37 °C with shock (180 rpm) until the OD 600 reached a value of 0.6.…”

Section: Methodsmentioning

confidence: 99%

Nanobody-Nanoluciferase Fusion Protein-Enabled Immunoassay for Ochratoxin A in Coffee with Enhanced Specificity and Sensitivity

Bao

Liu

Liao

et al. 2022

Toxins

View full text Add to dashboard Cite

Ochratoxin A (OTA), one of the best-known mycotoxins, causes problems concerning food safety with potential toxic effects in humans and animals. So, it is crucial to develop simple and sensitive methods for the detection of OTA. Herein, a nanoluciferase–nanobody fusion protein (Nb28-Nluc)-retaining antibody recognition and enzymatic activity was first prepared, which was then applied as a bifunctional tracer to construct a one-step bioluminescent enzyme-linked immunosorbent assay (BLEIA) for OTA in coffee samples. On the basis of Nb28-Nluc, the BLEIA can be completed with a one-step incubation and detection, with only a substrate replacement from 3,3′,5,5′-tetramethylbenzidine (TMB) to a Nluc assay reagent (Furimazine). Under the optimal experimental conditions, the proposed one-step BLEIA achieved a detection limit of 3.7 ng/mL (IC10) within 3 h. Moreover, the BLEIA method showed good repeatability and accuracy in the spike recovery experiments with recoveries of 83.88% to 120.23% and relative standard deviations (RSDs) of 5.2% to 24.7%, respectively. Particularly, the BLEIA displayed superior performances, such as fewer operations and more rapid and sensitive detection as compared with Nb28-based enzyme-linked immunosorbent assay. Therefore, the proposed one-step BLEIA has great potential for the sensitive and accurate screening of OTA in food samples.

show abstract

Section: Methodsmentioning

confidence: 99%

Nanobody-Nanoluciferase Fusion Protein-Enabled Immunoassay for Ochratoxin A in Coffee with Enhanced Specificity and Sensitivity

Bao

Liu

Liao

et al. 2022

Toxins

View full text Add to dashboard Cite

show abstract

“…Recently developed secondary nanobodies showed superior properties for cellular biology studies regarding penetrance, staining accuracy and, furthermore, secondary nanobodies can be premixed with primary antibodies to bypass the primary antibody animalspecies limitations (Sograte-Idrissi et al, 2020). Nanobodies can be fused to the Fc of conventional immunoglobulins and produced recombinantly, which complements nanobody binding capabilities with several technologies already available for monoclonal antibodies (Bao et al, 2021;Girt et al, 2021;Valenzuela Nieto et al, 2021) (Figure 1C).…”

Section: Nanobodies As Diagnostic Toolsmentioning

confidence: 99%

Nanobodies: COVID-19 and Future Perspectives

Nieto

Miranda-Chacón

Salinas-Rebolledo

et al. 2022

Front. Drug. Discov.

View full text Add to dashboard Cite

The COVID-19 pandemic has driven biotechnological developments to provide new and more effective tools for prophylaxis, diagnosis, and therapy. Historically, monoclonal antibodies have been valuable tools; however, the pandemic has shown some weaknesses, such as production limitations at a global scale. An alternative to conventional monoclonal antibodies are nanobodies, recombinant fragments of the variable region of single-domain antibodies derived mainly from the Camelidae family. Nanobodies have multiple characteristic benefits: they are small (15 KDa) and have remarkable refolding capability and unlimited possibilities for modifications due to their recombinant nature. Here, we review the application of nanobodies in diagnosis and treatment of SARS-CoV-2 infection.

show abstract

“…25 In recent years, reports on the development of multimeric nanobodies based on protein self-assembly have increased significantly due to simple operation and remarkable effects. Commonly used self-assembling proteins include right-handed coiled coil, 26 cartilage oligomeric matrix protein, 27 C4-binding protein, 28 and ferritin, 29 which can form a tetramer, pentamer, heptamer, and twenty-four mer, respectively. In our previous work, the OTA-specific nanobody (Nb28) was fused with the C4bpα C-terminal fragment (residues 541–597, C4bpα) to form a self-assembled heptamer nanobody.…”

Section: Introductionmentioning

confidence: 99%

Rapid and sensitive immunoassay for alpha-fetoprotein in serum by fabricating primary antibody–enzyme complexes using protein self-assembly

Liu

Peng

et al. 2023

Anal. Methods

Self Cite

View full text Add to dashboard Cite

show abstract

Nanobody multimerization strategy to enhance the sensitivity of competitive ELISA for detection of ochratoxin A in coffee samples

Cited by 36 publications

References 37 publications

Nanobody-Nanoluciferase Fusion Protein-Enabled Immunoassay for Ochratoxin A in Coffee with Enhanced Specificity and Sensitivity

Nanobody-Nanoluciferase Fusion Protein-Enabled Immunoassay for Ochratoxin A in Coffee with Enhanced Specificity and Sensitivity

Nanobodies: COVID-19 and Future Perspectives

Rapid and sensitive immunoassay for alpha-fetoprotein in serum by fabricating primary antibody–enzyme complexes using protein self-assembly

Contact Info

Product

Resources

About