Development of a streptavidin-bridged enhanced sandwich ELISA based on self-paired nanobodies for monitoring multiplex Salmonella serogroups

Ren, Yarong; Wei, Juan; Wang, Ye; Wang, Peng; Ji, Yanwei; Liu, Baoyuan; Wang, Jianlong; González‐Sapienza, Gualberto; Wang, Yanru

doi:10.1016/j.aca.2022.339705

Cited by 37 publications

(7 citation statements)

References 30 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…As the significant index to evaluate the sensitivity of the method, the limit of determination (LOD) was defined as the E. coli O157:H7 concentration corresponding to the OD value of the blank plus three standard deviations. 28 To evaluate the specificity of PdRu-ELISA, E. coli O157:H7 and other strains (E. coli, Listeria monocytogenes, Staphylococcus aureus, Vibrio parahaemolyticus, Salmonella, Bacillus subtilis, Salmonella enterica serovar, and Campylobacter jejuni) were detected at the same bacterial concentration (10 7 CFU/mL). PBS buffer was used as the negative control.…”

Section: ■ Materials and Methodsmentioning

confidence: 99%

See 1 more Smart Citation

A Versatile PdRu Bimetallic Nanoenzyme-Integrated Enzyme-Linked Immunosorbent Assay for Highly Sensitive Escherichia coli O157:H7 Detection

Wang,

Bu,

Cao

et al. 2023

Anal. Chem.

View full text Add to dashboard Cite

Nanozymes have drawn much attention as an enzyme mimetic with low cost and stability in enhancing analytical performance. Herein, a peroxidase-mimicking nanozyme-improved enzyme-linked immunosorbent assay (ELISA) was developed employing the bimetallic PdRu nanozyme to replace the natural enzymes as a catalytic carrier for the sensing of Escherichia coli O157:H7 (E. coli O157:H7). The PdRu nanozyme displayed ultrahigh catalytic activity, possessing a catalytic rate that was 5-fold higher than horseradish peroxidase (HRP). In addition, PdRu exhibited great biological affinity with antibody (affinity constant was about 6.75 × 1012 M) and high stability. All those advantages ensure the successful establishment and the construction of a novel colorimetric biosensor for E. coli O157:H7 detection. PdRu-based ELISA not only achieved an ultrasensitive detection sensitivity (8.7 × 102 CFU/mL) by approximately 288-fold as compared to the traditional HRP-based ELISA and also possessed satisfactory specificity and reproducibility (relative standard deviation (RSD) < 10%). Furthermore, the feasibility of PdRu-ELISA was further evaluated by detecting E. coli O157:H7 in actual samples with satisfactory recoveries, indicating its potential for applications in bioassays and clinical diagnostics.

show abstract

Section: ■ Materials and Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

A Versatile PdRu Bimetallic Nanoenzyme-Integrated Enzyme-Linked Immunosorbent Assay for Highly Sensitive Escherichia coli O157:H7 Detection

Wang,

Bu,

Cao

et al. 2023

Anal. Chem.

View full text Add to dashboard Cite

show abstract

“…Currently, the mainstream approach relies on recombinant modifications of nanobodies with their target-binding partners. This involves utilizing stationary systems such as biotin–streptavidin complexes through fusion of biotinylated nanobodies or employing biotin-binding proteins like rhizavidin, introducing lysine or cysteine residues tail at the C-terminus of nanobodies attempted for covalent attachment, using histidine tags for coordination interaction with Ni + -chelated surfaces and employing the SpyTag/SpyCatcher covalent ligation system . Additionally, several chemical modification methods, such as site-directed immobilization via sortase-mediated approaches, have also been explored .…”

Section: Introductionmentioning

confidence: 99%

Enhancing Oriented Immobilization Efficiency: A One-for-Two Organism-Bispecific Nanobody Scaffold for Highly Sensitive Detection of Foodborne Pathogens

Wang,

Zhang,

Wang

et al. 2023

Anal. Chem.

Self Cite

View full text Add to dashboard Cite

Nanobodies have gained widespread application in immunoassays. However, their small size presents a significant challenge in achieving effective immobilization and optimal sensitivity. Here, we present a novel “one-for-two”-oriented immobilization platform based on an organism-bispecific nanobody (O-BsNb) scaffold, enabling highly sensitive detection of two bacterial pathogens. Through genetic engineering, a bispecific nanobody (BsNb) was engineered, targeting Salmonella spp. and Vibrio parahaemolyticus. The O-BsNb scaffold allowed one nanobody to bind specifically to inactivated bacteria, forming an organism-oriented immobilization platform, while the other served as the capture antibody. Consequently, the O-BsNb bioscaffold-based ELISA (O-ELISA) for individual detection of S. enteritidis and V. parahaemolyticus was established. When compared to the sandwich ELISA utilizing passive immobilization of monovalent nanobodies, the O-ELISA exhibited a remarkable 13.4- and 13.7-fold improvement in LOD for S. enteritidis and V. parahaemolyticus, respectively, highlighting the enhanced immobilization efficacy of the O-ELISA. Furthermore, the feasibility and reproducibility of the assay in practical samples were meticulously evaluated, revealing exemplary performance in terms of recovery precision and assay stability. These findings demonstrate the significant potential of the O-ELISA platform for the sensitive detection of macromolecules, opening new avenues for efficient pathogen identification in foodborne safety and clinical diagnostics.

show abstract

“…To date, the gold-standard culture method is still the most reliable way to detect bacteria, but it is time-consuming and not suitable for on-site monitoring [ 6 , 7 ]. Other recommended methods, such as enzyme-linked immunosorbent assay (ELISA) and polymerase chain reaction (PCR), are time-saving, but ELISA has to screen specific antibodies and PCR requires expensive equipment [ 8 , 9 , 10 ]. Therefore, fast, accurate, low-cost and in-field detection techniques are in urgent need.…”

Section: Introductionmentioning

confidence: 99%

A Salmonella Microfluidic Chip Combining Non-Contact Eddy Heater and 3D Fan-Shaped Mixer with Recombinase Aided Amplification

Duan

Zhang

et al. 2022

Biosensors

View full text Add to dashboard Cite

Foodborne pathogenic bacteria have become a worldwide threat to human health, and rapid and sensitive bacterial detection methods are urgently needed. In this study, a facile microfluidic chip was developed and combined with recombinase-aided amplification (RAA) for rapid and sensitive detection of Salmonella typhimurium using a non-contact eddy heater for dynamic lysis of bacterial cells and a 3D-printed fan-shaped active mixer for continuous-flow mixing. First, the bacterial sample was injected into the chip to flow through the spiral channel coiling around an iron rod under an alternating electromagnetic field, resulting in the dynamic lysis of bacterial cells by this non-contact eddy heater to release their nucleic acids. After cooling to ~75 °C, these nucleic acids were continuous-flow mixed with magnetic silica beads using the fan-shaped mixer and captured in the separation chamber using a magnet. Finally, the captured nucleic acids were eluted by the eluent from the beads to flow into the detection chamber, followed by RAA detection of nucleic acids to determine the bacterial amount. Under the optimal conditions, this microfluidic chip was able to quantitatively detect Salmonella typhimurium from 1.1 × 102 to 1.1 × 105 CFU/mL in 40 min with a detection limit of 89 CFU/mL and might be prospective to offer a simple, low-cost, fast and specific bacterial detection technique for ensuring food safety.

show abstract

Development of a streptavidin-bridged enhanced sandwich ELISA based on self-paired nanobodies for monitoring multiplex Salmonella serogroups

Cited by 37 publications

References 30 publications

A Versatile PdRu Bimetallic Nanoenzyme-Integrated Enzyme-Linked Immunosorbent Assay for Highly Sensitive Escherichia coli O157:H7 Detection

A Versatile PdRu Bimetallic Nanoenzyme-Integrated Enzyme-Linked Immunosorbent Assay for Highly Sensitive Escherichia coli O157:H7 Detection

Enhancing Oriented Immobilization Efficiency: A One-for-Two Organism-Bispecific Nanobody Scaffold for Highly Sensitive Detection of Foodborne Pathogens

A Salmonella Microfluidic Chip Combining Non-Contact Eddy Heater and 3D Fan-Shaped Mixer with Recombinase Aided Amplification

Contact Info

Product

Resources

About