Preparation of Specific Nanobodies and Their Application in the Rapid Detection of Nodularin-R in Water Samples

Nanobodies have several advantages, including great stability, sensibility, and ease of production; therefore, they have become important tools in immunoassays for chemical contaminants. In this manuscript, nanobodies for the detection of the toxin Nodularin-r (NOD-R), a secondary metabolite of cyanobacteria that could cause a safety risk for drinks and food for its strong hepatotoxicity, were for the first time selected from an immunized Bactrian camel VHH phage display library. Then, a sensitive indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) for NOD-R, based on the nanobody N56 with great thermostability and organic solvent tolerance, was established under optimized conditions. The results showed that the limit of detection for NOD-R was 0.67 µg/L, and the average spike recovery rate was between 84.0 and 118.3%. Moreover, the ic-ELISA method was validated with spiked water sample and confirmed by UPLC–MS/MS, which indicated that the ic-ELISA established in this work is a reproducible detection assay for nodularin residues in water samples.

Download Full-text

Detection of bisphenol A in water samples using ELISA determination method

Water Science & Technology Water Supply ◽

10.2166/ws.2011.008 ◽

2011 ◽

Vol 11 (1) ◽

pp. 55-60 ◽

Cited By ~ 4

Author(s):

J. Zheng ◽

S. Q. Zhao ◽

X. T. Xu ◽

K. Zhang

Keyword(s):

Drinking Water ◽

Bisphenol A ◽

Water Samples ◽

Limit Of Detection ◽

Industrial Effluent ◽

Cross Reactivity ◽

Enzyme Linked Immunosorbent Assay ◽

Uv Spectrophotometry ◽

Inhibition Concentration

In order to study whether bisphenol A (BPA) can pass into drinking water from polycarbonate barrel and exist in the river and industrial effluent the indirect competitive enzyme-linked immunosorbent assay (ELISA) for the determination of BPA was established. The results presented an inhibition concentration at 50% absorbance (IC50) of 0.123 mg L−1, and the limit of detection (LOD) is 9.934 μg L−1. The specificity of antiserum was proved well because the cross-reactivity with benzene, tert-butylbenzene, hydroquinone and o-hydroxybenzoic acid were found lower than 0.01%, except phenol was 0.26%. The method was found to be reliable and repeatable. It was used for monitoring the concentration of BPA in the barreled drinking water. The results confirmed BPA can pass into barreled drinking water from the polycarbonate barrel and concentration increased as days went on. A certain content of BPA was found in industrial effluent. The results of ELISA were consistent with the results of UV spectrophotometry. BPA could not be found in the water samples obtained from Zhujiang River. The established method shows specific recognition of BPA and could be applied in detection of environmental BPA.

Download Full-text

Paper-Based Analytical Device for Rapid Naked-Eye Detection of Sulfide in Water Samples

Journal of the chemical society of pakistan ◽

10.52568/000681 ◽

2020 ◽

Vol 42 (5) ◽

pp. 696-696

Author(s):

Lingzhi Zhao Lingzhi Zhao

Keyword(s):

Water Samples ◽

Limit Of Detection ◽

Rapid Determination ◽

Tap Water ◽

Low Cost ◽

Color Intensity ◽

Detection Zone ◽

Simulated Waste ◽

Optimized Conditions ◽

Analytical Device

This paper describes the development of a low-cost paper-based colorimetric analytical device for the accurate and rapid determination of sulfide. Under optimized conditions, Na2S was dropped in the uptake zone I and the probe in the uptake zone II, they converged to the detection zone via capillary force and formed an intense pink resorufin. Sulfide can be quantified based on the average color intensity values of the product “free resorufin”. The color intensity is recorded using a camera phone, and quantification was made using Adobe Photoshop. The as-developed analytical device detected sulfide in the range of 5–400 μM (R2 = 0.982) with the limit of detection (LOD) 1 μM, and was successfully applied in sulfide assay in spiked water samples including tap water and simulated waste water. Colorimetric results from the proposed paper-based colorimetric analytical device were consistent with that from methylene blue (MB) method.

Download Full-text

Design of Novel Haptens and Development of Monoclonal Antibody-Based Immunoassays for the Simultaneous Detection of Tylosin and Tilmicosin in Milk and Water Samples

Biomolecules ◽

10.3390/biom9120770 ◽

2019 ◽

Vol 9 (12) ◽

pp. 770 ◽

Cited By ~ 2

Author(s):

Jian-Xin Huang ◽

Chan-Yuan Yao ◽

Jin-Yi Yang ◽

Zhen-Feng Li ◽

Fan He ◽

...

Keyword(s):

Monoclonal Antibody ◽

Water Samples ◽

Simultaneous Detection ◽

Aldehyde Group ◽

Cross Reactivity ◽

Side Chain ◽

Enzyme Linked Immunosorbent Assay ◽

Inhibition Concentration ◽

Ic50 Values ◽

Optimized Conditions

In this work, a monoclonal antibody-based indirect competitive enzyme-linked immunosorbent assay (icELISA) was established to detect tylosin and tilmicosin in milk and water samples. A sensitive and specific monoclonal antibody was prepared by rational designed hapten, which was achieved by directly oxidizing the aldehyde group on the side chain of tylosin to the carboxyl group. Under the optimized conditions, the linear range of icELISA for tylosin and tilmicosin were 1.3 to 17.7 ng/mL and 2.0 to 47.4 ng/mL, with half-maximal inhibition concentration (IC50) values of 4.7 and 9.6 ng/mL, respectively. The cross-reactivity with other analogues of icELISA was less than 0.1%. The average recoveries of icELISA for tylosin and tilmicosin ranged from 76.4% to 109.5% in milk and water samples. Besides, the detection results of icELISA showed good correlations with HPLC-MS/MS. The proposed icELISA was satisfied for rapid and specific screening of tylosin and tilmicosin residues in milk and water samples.

Download Full-text

Spacer layer engineering for ultrasensitive Hg(II) detection on surface plasmon-coupled emission platform

Nanotechnology Reviews ◽

10.1515/ntrev-2017-0124 ◽

2017 ◽

Vol 6 (4) ◽

pp. 331-338

Author(s):

Pradeep Kumar Badiya ◽

Tejkiran Pindi Jayakumar ◽

Venkatesh Srinivasan ◽

Sai Sathish Ramamurthy

Keyword(s):

Surface Plasmon ◽

Water Samples ◽

Limit Of Detection ◽

Water Security ◽

Diagnostic Tools ◽

Mercury Ions ◽

Spacer Layer ◽

Analyte Detection ◽

Field Device ◽

First Time

AbstractIn this work, we demonstrate for the first time the ultrasensitive detection of Hg2+ ions with femtomolar sensitivity in water samples with the use of the surface plasmon-coupled emission (SPCE) platform. The use of portable network diagnostic tools for water security and integrated water shed management is a topic of recent research interest. In this context, the current study explores Hg2+ monitoring using a rhodamine-6G (Rh6G) derivative bearing a monothiospirolactone mounted onto a SPCE substrate. Thus far, the limit of detection for mercury ions by the conventional fluorescence technique has been 0.15 nm. However, we have achieved 1 fm Hg2+ detection using silver nanoparticle-based spacer layer engineering on an SPCE sensor chip. Using this technology, a field device can be fabricated for rapid, ultrasensitive, multi-analyte detection (of contaminants) in water samples.

Download Full-text

Carrier-Free Immobilization of α-Galactosidase as Nano-Biocatalysts for Synthesizing Prebiotic α-Galacto-Oligosaccharides

Molecules ◽

10.3390/molecules26051248 ◽

2021 ◽

Vol 26 (5) ◽

pp. 1248

Author(s):

Yan Liu ◽

Jingyi Yang ◽

Ke Wang ◽

Feiyu Duan ◽

Lili Lu

Keyword(s):

Ammonium Sulfate ◽

Industrial Applications ◽

Solvent Tolerance ◽

Cross Linking ◽

Organic Solvent Tolerance ◽

Regular Structures ◽

Carrier Free ◽

High Precipitation ◽

First Time ◽

Scanning Electron

α-Galacto-oligosaccharides (α-GOSs) have great functions as prebiotics and therapeutics. This work established the method of batch synthesis of α-GOSs by immobilized α-galactosidase for the first time, laying a foundation for industrial applications in the future. The α-galactosidase from Aspergillus niger L63 was immobilized as cross-linked enzyme aggregates (CLEAs) nano-biocatalyst through enzyme precipitating and cross-linking steps without using carriers. Among the tested agents, the ammonium sulfate showed high precipitation efficacy and induced regular structures of α-galactosidase CLEAs (Aga-CLEAs) that had been analyzed by scanning electron microscopy and Fourier-transform infrared spectroscopy. Through optimization by response surface methodology, the ammonium sulfate-induced Aga-CLEAs achieved a high activity recovery of around 90% at 0.55 U/mL of enzymes and 36.43 mM glutaraldehyde with cross-linking for 1.71 h. Aga-CLEAs showed increased thermal stability and organic solvent tolerance. The storage ability was also improved since it maintained 74.5% activity after storing at 4 °C for three months, significantly higher than that of the free enzyme (21.6%). Moreover, Aga-CLEAs exhibited excellent reusability in the α-GOSs synthesis from galactose, retaining above 66% of enzyme activity after 10 batch reactions, with product yields all above 30%.

Download Full-text

Vortex-assisted liquid–liquid microextraction combined with spectrophotometry for the determination of trace nitrite in water samples

Water Science & Technology Water Supply ◽

10.2166/ws.2017.010 ◽

2017 ◽

Vol 17 (5) ◽

pp. 1225-1231 ◽

Cited By ~ 1

Author(s):

Yang Jiao ◽

Jianping Yu ◽

Yaling Yang

Keyword(s):

Organic Solvent ◽

Water Samples ◽

Limit Of Detection ◽

Environmental Water ◽

Cationic Dye ◽

Optimum Conditions ◽

Relative Standard ◽

Optimized Conditions ◽

Liquid Microextraction

A vortex-assisted liquid–liquid microextraction (VALLME) method using isooctanol as extractant followed by spectrophotometry was developed for the extraction and determination of trace nitrite in water samples. The method is based on selective ion-pairing complex (I3− MG+) formation of triiodideanion I3− with cationic dye malachite green (MG) at pH 3.0, and its subsequent extraction in an organic solvent. The extracted organic solvent-rich phase is diluted with methanol, and its absorbance is measured against an analyte blank at 630 nm. The variables affecting VALLME efficiency were investigated, and a set of optimized conditions was obtained. Under the optimum conditions, the linear range of nitrite was from 1.0 to 100 ng mL−1. The relative standard deviations (n = 10) were 2.1–3.9% and the limit of detection was 0.5 ng mL−1 and was successfully applied to the determination of nitrite in environmental water.

Download Full-text

A rapid magnetic bead-based immunoassay for sensitive determination of diclofenac

Analytical and Bioanalytical Chemistry ◽

10.1007/s00216-021-03778-7 ◽

2021 ◽

Author(s):

Alexander Ecke ◽

Tanja Westphalen ◽

Jane Hornung ◽

Michael Voetz ◽

Rudolf J. Schneider

Keyword(s):

Water Samples ◽

Magnetic Beads ◽

Limit Of Detection ◽

Cost Effective ◽

Magnetic Bead ◽

Enzyme Linked Immunosorbent Assay ◽

Reference Analysis ◽

Immunochemical Method ◽

Synthetic Strategy

Abstract Increasing contamination of environmental waters with pharmaceuticals represents an emerging threat for the drinking water quality and safety. In this regard, fast and reliable analytical methods are required to allow quick countermeasures in case of contamination. Here, we report the development of a magnetic bead-based immunoassay (MBBA) for the fast and cost-effective determination of the analgesic diclofenac (DCF) in water samples, based on diclofenac-coupled magnetic beads and a robust monoclonal anti-DCF antibody. A novel synthetic strategy for preparation of the beads resulted in an assay that enabled for the determination of diclofenac with a significantly lower limit of detection (400 ng/L) than the respective enzyme-linked immunosorbent assay (ELISA). With shorter incubation times and only one manual washing step required, the assay demands for remarkably shorter time to result (< 45 min) and less equipment than ELISA. Evaluation of assay precision and accuracy with a series of spiked water samples yielded results with low to moderate intra- and inter-assay variations and in good agreement with LC–MS/MS reference analysis. The assay principle can be transferred to other, e.g., microfluidic, formats, as well as applied to other analytes and may replace ELISA as the standard immunochemical method. Graphical abstract

Download Full-text

Nanobody-Alkaline Phosphatase Fusion Protein-Based Enzyme-Linked Immunosorbent Assay for One-Step Detection of Ochratoxin A in Rice

Sensors ◽

10.3390/s18114044 ◽

2018 ◽

Vol 18 (11) ◽

pp. 4044 ◽

Cited By ~ 6

Author(s):

Zhichang Sun ◽

Xuerou Wang ◽

Qi Chen ◽

Yonghuan Yun ◽

Zongwen Tang ◽

...

Keyword(s):

Alkaline Phosphatase ◽

Fusion Protein ◽

Ochratoxin A ◽

Immunosorbent Assay ◽

Limit Of Detection ◽

Binding Capacity ◽

Cross Reactivity ◽

Solvent Tolerance ◽

Enzyme Linked Immunosorbent Assay ◽

One Step

Ochratoxin A (OTA) has become one a focus of public concern because of its multiple toxic effects and widespread contamination. To monitor OTA in rice, a sensitive, selective, and one-step enzyme-linked immunosorbent assay (ELISA) using a nanobody-alkaline phosphatase fusion protein (Nb28-AP) was developed. The Nb28-AP was produced by auto-induction expression and retained an intact antigen-binding capacity and enzymatic activity. It exhibited high thermal stability and organic solvent tolerance. Under the optimal conditions, the developed assay for OTA could be finished in 20 min with a half maximal inhibitory concentration of 0.57 ng mL−1 and a limit of detection of 0.059 ng mL−1, which was 1.1 times and 2.7 times lower than that of the unfused Nb28-based ELISA. The Nb28-AP exhibited a low cross-reactivity (CR) with ochratoxin B (0.92%) and ochratoxin C (6.2%), and an ignorable CR (<0.10%) with other mycotoxins. The developed Nb-AP-based one-step ELISA was validated and compared with a liquid chromatography-tandem mass spectrometry method. The results show the reliability of Nb-AP-based one-step ELISA for the detection of OTA in rice.

Download Full-text

Enzyme-linked immunosorbent assay for triclocarban in aquatic environments

Water Science & Technology ◽

10.2166/wst.2015.366 ◽

2015 ◽

Vol 72 (10) ◽

pp. 1682-1691 ◽

Cited By ~ 3

Author(s):

Kun Zeng ◽

Yanmin Zou ◽

Jianxia Liu ◽

Wei Wei ◽

Meng Zhang ◽

...

Keyword(s):

Monoclonal Antibody ◽

Immunosorbent Assay ◽

Limit Of Detection ◽

Enzyme Linked Immunosorbent Assay ◽

Aquatic Environments ◽

Satisfactory Accuracy ◽

Good Linear ◽

High Affinity ◽

Inhibition Concentration ◽

Optimized Conditions

A sensitive, competitive indirect enzyme-linked immunosorbent assay (ELISA) was developed for the detection of triclocarban (TCC) in waters and sediments. Haptens were synthesized by derivatizing the paraposition of a phenyl moiety of TCC. The synthesized hapten was then coupled to bovine thyroglobulin to be used as an immunogen, based on which, a high affinity monoclonal antibody 4D5 was produced with the hybridoma technique. Under the optimized conditions, using the monoclonal antibody, excellent performances of the assay were obtained: satisfactory sensitivity (IC50 (50% inhibition concentration) value, 0.43 ng/mL; limit of detection, 0.05 ng/mL); good linear range (0.05–10 ng/mL); and satisfactory accuracy (recoveries 70.7–107% in waters; 74.8–98.3% in sediments). Furthermore, TCC was found with the concentration ranging from not detected to 422.12 ng/L in waters and from 6.68 ng/g to 78.67 ng/g in sediments in Yunliang River, Ancient Canal and Hongqiao Port in Zhenjiang City. In conclusion ELISA could be applied for monitoring TCC in aquatic environments.

Download Full-text

Integration of Enzyme-Linked Immunosorbent Assay with Conventional Chromatographic Procedures for Quantitation of Aflatoxin in Individual Cotton Bolls, Seeds, and Seed Sections

Journal of AOAC INTERNATIONAL ◽

10.1093/jaoac/73.4.581 ◽

1990 ◽

Vol 73 (4) ◽

pp. 581-584 ◽

Cited By ~ 1

Author(s):

Louise S Lee ◽

James H Wall ◽

Peter J Cotty ◽

Paul Bayman

Keyword(s):

Immunosorbent Assay ◽

Limit Of Detection ◽

Rapid Assessment ◽

Time Integration ◽

Extraction Procedure ◽

Distribution Patterns ◽

Enzyme Linked Immunosorbent Assay ◽

Large Numbers ◽

First Time ◽

Cotton Bolls

Abstract Integration of an enzyme-linked Immunosorbent assay (ELISA) with conventional chromatographic methods proved the versatility of ELISA as a research tool and allowed for rapid assessment of aflatoxin In Individual cottonseeds, parts of cottonseed, and composite samples of seeds taken from Individual cotton bolls. Aqueous acetone was substituted for methanol In the extraction procedure prescribed by ELISA. The substitution allowed the use of a common extract for all analytical methods. An aliquot of the extract was used to screen samples by ELISA. Negative samples were Identified, and toxin levels between 1 and 70 ng/g were quantltated by ELISA. Samples with toxin levels beyond the upper limit of detection by ELISA were then subjected to more time-consuming conventional cleanup prior to quantitation by liquid chromatography (LC) or thin-layer chromatography (TLC). Toxin levels detected by LC or TLC ranged from 100 to 845 000 ng/g sample. The screen by ELISA detected large numbers of toxin-negative cotton bolls or Individual seeds in minimum analysis time. The combination of techniques verified the presence of seed with no toxin adjacent to toxincontaining seed In the same lock. Toxin-negative portions of Individual seed with high toxin In another portion were Identified for the first time. Integration of techniques provided needed Information on distribution patterns of aflatoxin In cotton so that preventive measures can be developed.

Download Full-text