Pitfalls in the Immunochemical Determination of β‑Lactam Antibiotics in Water

Contamination of waters with pharmaceuticals is an alarming problem as it may support the evolution of antimicrobial resistance. Therefore, fast and cost-effective analytical methods for potential on-site analysis are desired in order to control the water quality and assure the safety of its use as a source of drinking water. Antibody-based methods, such as the enzyme-linked immunosorbent assay (ELISA), can be helpful in this regard but can also have certain pitfalls in store, depending on the analyte. As shown here for the class of β-lactam antibiotics, hydrolysis of the β‑lactam ring is a key factor in the immunochemical analysis as it influences antibody recognition. With the antibody used in this study, the limit of detection (LOD) in the immunoassay could be significantly reduced by hydrolysis for the five tested penicillins, with the lowest LOD for carbenicillin (0.2 nmol/L) and the greatest impact on penicillins G and V (reduction by 85%). In addition to enhanced quantification, our strategy also provides access to information about the degree of hydrolysis in water samples as shown for the most abundant penicillin amoxicillin.

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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

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Immobilization-Free Electrochemical Sensor Coupled with a Graphene-Oxide-Based Aptasensor for Glycated Albumin Detection

Biosensors ◽

10.3390/bios11030085 ◽

2021 ◽

Vol 11 (3) ◽

pp. 85

Author(s):

Wassa Waiwinya ◽

Thitirat Putnin ◽

Dechnarong Pimalai ◽

Wireeya Chawjiraphan ◽

Nuankanya Sathirapongsasuti ◽

...

Keyword(s):

Graphene Oxide ◽

Electrochemical Sensor ◽

Detection Method ◽

Limit Of Detection ◽

Glycated Albumin ◽

Cost Effective ◽

Logarithmic Scale ◽

Electrochemical Aptasensor ◽

Alternative Approach

An immobilization-free electrochemical sensor coupled with a graphene oxide (GO)-based aptasensor was developed for glycated human serum albumin (GHSA) detection. The concentration of GHSA was monitored by measuring the electrochemical response of free GO and aptamer-bound GO in the presence of glycated albumin; their currents served as the analytical signals. The electrochemical aptasensor exhibited good performance with a base-10 logarithmic scale. The calibration curve was achieved in the range of 0.01–50 µg/mL. The limit of detection (LOD) was 8.70 ng/mL. The developed method was considered a one-drop measurement process because a fabrication step and the probe-immobilization process were not required. This simple sensor offers a cost-effective, rapid, and sensitive detection method, and could be an alternative approach for determination of GHSA levels.

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Characterization and validation of fragment antigen-binding (Fab) antibody-based immunoassay for deoxymiroestrol, a potent phytoestrogen from Pueraria candollei

10.22541/au.161270515.50311593/v1 ◽

2021 ◽

Author(s):

Worapol Sae-foo ◽

Supaluk Krittanai ◽

Wipawee Juengsanguanpornsuk ◽

Gorawit Yusakul ◽

Tharita Kitisripanya ◽

...

Keyword(s):

Quantitative Method ◽

Estrogenic Activity ◽

Limit Of Detection ◽

Hormone Replacement ◽

Enzyme Linked Immunosorbent Assay ◽

Antigen Binding ◽

Pueraria Candollei ◽

Specific Determination ◽

Fragment Antigen Binding

Deoxymiroestrol is the most potent phytoestrogen in chromenes group that has been found in Pueraria candollei, Thai name known as Kwao Krua Khao. Several studies reported estrogenic activity of P. candollei in order to using as hormone replacement therapy for postmenopausal women. Previously, specific determination of deoxymiroestrol content by enzyme-linked immunosorbent assay (ELISA) using polyclonal antibody (pAb) have been reported. However, production of pAb has limitation and variability from different batches. Therefore, in this study, we established quantitative method for determination of deoxymiroestrol using fragment antigen-binding (Fab) antibody-based immunoassay. The developed immunoassay has specificity to deoxymiroestrol with a calibration range of 15.6-1000 ng mL-1. Precision including intra-assay and inter-assay are 1.48-7.11 and 0.58-9.31%, respectively. Accuracy of the assay showed in recovery between 99.77-101.61% when spike deoxymiroestrol standard into the samples. The limit of detection (LOD) is 30.80 ng mL-1. Comparation antibody-based immunoassay for determination of deoxymiroestrol using Fab with pAb was represented consistency (R2 = 0.9807) when analysis roots bark of Pueraria candollei from difference areas. Therefore, this development assay can apply to determine deoxymiroestrol content in the plant samples.

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Proposal of De Novo Antigen Test for COVID-19: Ultrasensitive Detection of Spike Proteins of SARS-CoV-2

Diagnostics ◽

10.3390/diagnostics10080594 ◽

2020 ◽

Vol 10 (8) ◽

pp. 594 ◽

Cited By ~ 8

Author(s):

Yuta Kyosei ◽

Mayuri Namba ◽

Sou Yamura ◽

Rikiya Takeuchi ◽

Noriko Aoki ◽

...

Keyword(s):

De Novo ◽

Limit Of Detection ◽

False Negative ◽

Cost Effective ◽

Detection Sensitivity ◽

Enzyme Linked Immunosorbent Assay ◽

Antigen Test ◽

Virus Rna ◽

Antigen Tests ◽

Spike Proteins

Polymerase chain reaction (PCR)-based antigen tests are technically difficult, time-consuming, and expensive, and may produce false negative results requiring follow-up confirmation with computed tomography. The global coronavirus disease 2019 (COVID-19) pandemic has increased the demand for accurate, easy-to-use, rapid, and cost-effective antigen tests for clinical application. We propose a de novo antigen test for diagnosing COVID-19 using the combination of sandwich enzyme-linked immunosorbent assay and thio-nicotinamide adenine dinucleotide (thio-NAD) cycling. Our test takes advantage of the spike proteins specific to the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus. The limit of detection of our test was 2.3 × 10−18 moles/assay. If the virus has ~25 spike proteins on its surface, our method should detect on the order of 10−20 moles of virus/assay, corresponding to ~104 copies of the virus RNA/assay. The detection sensitivity approaches that of PCR-based assays because the average virus RNA load used for PCR-based assays is ~105 copies per oro- or naso-pharyngeal swab specimen. To our knowledge, this is the first ultrasensitive antigen test for SARS-CoV-2 spike proteins that can be performed with an easy-to-use microplate reader. Sufficient sensitivity can be achieved within 10 min of thio-NAD cycling. Our antigen test allows for rapid, cost-effective, specific, ultrasensitive, and simultaneous multiple measurements of SARS-CoV-2, and has broad application for the diagnosis for COVID-19.

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Determination of carcinogenic herbicides in milk samples using green non-ionic silicone surfactant of cloud point extraction and spectrophotometry

Royal Society Open Science ◽

10.1098/rsos.171500 ◽

2018 ◽

Vol 5 (4) ◽

pp. 171500 ◽

Cited By ~ 1

Author(s):

N. I. Mohd ◽

N. N. M. Zain ◽

M. Raoov ◽

S. Mohamad

Keyword(s):

Cloud Point ◽

Cloud Point Extraction ◽

Limit Of Detection ◽

Cost Effective ◽

Silicone Surfactant ◽

Extraction Solvent ◽

Milk Samples ◽

Relative Standard ◽

Limit Of Quantitation

A new cloud point methodology was successfully used for the extraction of carcinogenic pesticides in milk samples as a prior step to their determination by spectrophotometry. In this work, non-ionic silicone surfactant, also known as 3-(3-hydroxypropyl-heptatrimethylxyloxane), was chosen as a green extraction solvent because of its structure and properties. The effect of different parameters, such as the type of surfactant, concentration and volume of surfactant, pH, salt, temperature, incubation time and water content on the cloud point extraction of carcinogenic pesticides such as atrazine and propazine, was studied in detail and a set of optimum conditions was established. A good correlation coefficient ( R 2 ) in the range of 0.991–0.997 for all calibration curves was obtained. The limit of detection was 1.06 µg l −1 (atrazine) and 1.22 µg l −1 (propazine), and the limit of quantitation was 3.54 µg l −1 (atrazine) and 4.07 µg l −1 (propazine). Satisfactory recoveries in the range of 81–108% were determined in milk samples at 5 and 1000 µg l −1 , respectively, with low relative standard deviation, n = 3 of 0.301–7.45% in milk matrices. The proposed method is very convenient, rapid, cost-effective and environmentally friendly for food analysis.

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A Monoclonal Antibody-Based Enzyme-Linked Immunosorbent Assay for Determination of Homoharringtonine

Planta Medica ◽

10.1055/a-0578-8689 ◽

2018 ◽

Vol 84 (14) ◽

pp. 1038-1044 ◽

Cited By ~ 3

Author(s):

Benyakan Pongkitwitoon ◽

Seiichi Sakamoto ◽

Rika Nagamitsu ◽

Waraporn Putalun ◽

Hiroyuki Tanaka ◽

...

Keyword(s):

Monoclonal Antibody ◽

Immunosorbent Assay ◽

Myeloid Leukemia ◽

High Performance ◽

Sodium Periodate ◽

Limit Of Detection ◽

Enzyme Linked Immunosorbent Assay ◽

Study Support ◽

Mediated Oxidation

AbstractHomoharringtonine (HHT), also known as omacetaxine, is a natural compound found in the genus Cephalotaxus and is a promising pharmaceutical drug used for the treatment of chronic or accelerated phase chronic myeloid leukemia. As a tool for the quantitative determination of HHT, a specific monoclonal antibody against HHT (MAb 6A1) was generated by conjugates prepared via sodium periodate-mediated oxidation. The developed indirect competitive enzyme-linked immunosorbent assay (icELISA) using MAb 6A1 was found to be highly specific and sensitive with a limit of detection for HHT of 48.8 ng/mL. Validation assays to evaluate precision and accuracy of the method were conducted by the use of intra- and inter-assay analysis, recovery test, and comparison analysis between the amounts of HHT determined by ELISA and high-performance liquid chromatography. These results revealed that the established icELISA using MAb 6A1 is specific, sensitive, and reliable enough to be applied to the qualitative analysis for HHT. Furthermore, the results of this study support the usefulness of sodium periodate as a reagent for the conjugation between Cephalotaxus alkaloids and proteins for producing specific antibodies.

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Potential methodologies and strategies for the rapid assessment of feed-grain quality

Australian Journal of Agricultural Research ◽

10.1071/ar98167 ◽

1999 ◽

Vol 50 (5) ◽

pp. 789 ◽

Cited By ~ 11

Author(s):

C. W. Wrigley

Keyword(s):

Grain Quality ◽

Near Infrared ◽

Animal Feed ◽

Rapid Assessment ◽

Cost Effective ◽

Beta Glucan ◽

Site Analysis ◽

Direct Indication ◽

Systematic Strategy

The efficient use of grains for animal feed requires the use of analytical methods that can provide rapid indications of the suitability of the grain for animal nutrition. Ideally, these methods need to be applied at the site of grain receival within the tight time and cost confines of grain delivery. In addition, methods are needed in plant breeding to efficiently screen for target aspects of feed-grain quality to facilitate the development of genotypes with improved nutritional quality. This review describes a range of techniques that can fulfil these analytical requirements. These include visual examination of grain samples for species identification and for recognition of defects and contaminants. This long-standing approach is rapid, but it is subjective and dependent on the expertise of the operator. The newer technology of image analysis offers the prospect of providing similar information automatically and quantitatively, without the risk of operator bias. Near-infrared (NIR) analysis is already in general use for grain analysis at many receival depots in wheat-growing countries, mainly for the determination of moisture and protein content. There are promising indications that NIR can be extended to the determination of many other aspects of grain composition, both the positive aspects that contribute to feed quality, as well as components such as beta-glucan content that have negative contributions for non-ruminants. Furthermore, NIR is being developed to provide a direct indication of metabolisable energy for a range of grain types. Whereas NIR is primarily suited to the determination of quantitatively major components of the grain, without the requirement of significant sample preparation, immunological analyses are appropriate for the determination of specific and minor components, such as mycotoxins, lectins, alkaloids, and pesticide residues. These and other methods, suited for on-site analysis, need to be combined with effective sampling to ensure that the results of testing are representative of the whole of the grain consignment, and also integrated into a systematic strategy to ensure cost-effective testing.

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Development of an Enzyme-Linked Immunosorbent Assay for Determination Of Miroestrol Using an Anti-miroestrol Monoclonal Antibody

Planta Medica ◽

10.1055/s-0043-102689 ◽

2017 ◽

Vol 83 (10) ◽

pp. 855-861 ◽

Cited By ~ 7

Author(s):

Tharita Kitisripanya ◽

Supaluk Krittanai ◽

Orapin Udomsin ◽

Kamonthip Jutathis ◽

Jukrapun Komaikul ◽

...

Keyword(s):

Monoclonal Antibody ◽

Estrogenic Activity ◽

Limit Of Detection ◽

Simple Procedure ◽

Enzyme Linked Immunosorbent Assay ◽

Efficacy And Safety ◽

Limit Of Quantitation ◽

Estrogenic Action ◽

White Kwao Krua

AbstractMiroestrol is a chromene with potent estrogenic activity present in Pueraria candollei, commonly known as White Kwao Krua. Although this compound is only present in low amounts in the plant, it plays an important role in the estrogenic action of P. candollei products. As a tool for further studies about the efficacy and safety of P. candollei as a phytoestrogenic supplement, we generated a novel monoclonal antibody against miroestrol. This anti-miroestrol monoclonal antibody was used to develop an immunoassay for the determination of miroestrol content, which can be used for quality control purposes of P. candollei. The developed ELISA against miroestrol has a calibration range of 10–780 ng/mL miroestrol, a limit of detection of 3.5 ng/mL, and a limit of quantitation of 12.2 ng/mL. According to the validation analysis, the established ELISA is precise, accurate, specific, and sensitive for miroestrol detection in plants. Furthermore, the anti-miroestrol monoclonal antibody was used to prepare an immunoaffinity column for the isolation of miroestrol from the tuberous root of P. candollei. The column provides a simple procedure for miroestrol isolation, with a capacity of 3.91 µg of miroestrol per 1 mL of immunogel.

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Graphitic-Phase C3N4 Nanosheets Combined with MnO2 Nanosheets for Sensitive Fluorescence Quenching Detection of Parathion-Methyl

10.21203/rs.3.rs-210957/v1 ◽

2021 ◽

Author(s):

Bicheng Liu ◽

Sihao Wu ◽

Zoujun Peng ◽

Jiahan Rui ◽

Ping Qiu

Keyword(s):

Fluorescence Quenching ◽

Limit Of Detection ◽

Organophosphorus Pesticides ◽

Cost Effective ◽

Inner Filter Effect ◽

Enzymatic Hydrolysate ◽

Analytical Instruments ◽

Dimethyl Phosphate ◽

User Friendly

Abstract In this study, we have developed a sensitive approach to measure organophosphorus pesticides (OPs) using graphitic-phase C3N4 nanosheets (g-C3N4) combined with a nanomaterial-based quencher MnO2 nanosheets (MnO2 NS). Because MnO2 NS could quench the fluorescence of g-C3N4 via the inner-filter effect (IFE), the enzymatic hydrolysate (thiocholine, TCh) can efficiently trigger the decomposition of MnO2 nanosheets in the presence of acetylcholinesterase (AChE) and acetylthiocholine, resulting in the fluorescence recovery of g-C3N4. OPs, as inhibitors for AChE activity, can prevent the generation of TCh and decomposition of MnO2 nanosheets, accompanied by fluorescence quenching again. So the AChE-ATCh-MnO2-g-C3N4 system can be utilized to detect OPs quantitatively based on the g-C3N4 fluorescence. Under the optimum conditions, the linear range for the determination of parathion-methyl (PM) and 2,2-dichlorovinyl dimethyl phosphate (DDVP) were found in the range of 0.1-2.1 ng/mL with a limit of detection of 0.069 ng/mL, and 0.5-16 ng/mL with a limit of detection of 0.069 ng/mL, respectively. Finally, this method was exploited for the monitoring of PM in real samples. The advantages of the assay are user-friendly, easy-to-ease, cost-effective compared to sophisticated analytical instruments.

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Vortex-Assisted Dispersive Liquid–Liquid Microextraction Coupled with Deproteinization for Determination of Nateglinide in Human Plasma Using HPLC/UV

Journal of Chromatographic Science ◽

10.1093/chromsci/bmaa096 ◽

2020 ◽

Author(s):

Mohamed A Hammad ◽

Amira H Kamal ◽

Reham E Kannouma ◽

Fotouh R Mansour

Keyword(s):

Human Plasma ◽

High Performance ◽

Limit Of Detection ◽

Signal To Noise Ratio ◽

Cost Effective ◽

Salt Addition ◽

Relative Standard ◽

Dispersive Liquid Liquid Microextraction ◽

Liquid Microextraction

Abstract A validated method for preconcentration and determination of nateglinide in plasma was developed using vortex-assisted dispersive liquid–liquid microextraction. Different variables that affect extraction efficiency were studied and optimized, including type and volume of extractant, type and volume of disperser, pH of diluent, salt addition effect, centrifugation and vortex time. Nateglinide was extracted using 30 μL of 1-octanol as an extractant and 200 μL of methanol as a disperser. The enrichment factor reached 330 under the optimum conditions. High-performance liquid chromatography/ultraviolet was used for detection using phosphate buffer (pH 2.5, 10 mM): acetonitrile (45:55, v/v) as a mobile phase at a flow rate of 1 mL/min. The method was linear over the range of 50–20,000 ng/mL with a limit of detection of 15 ng/mL (signal-to-noise ratio = 3). Intra- and inter-day precision had %relative standard deviation <6% (n = 3) and the %recoveries were found to be between 102.5 and 105.9%. The proposed method is simple, sensitive, eco-friendly, cost-effective and powerful for microextraction of nateglinide from human plasma samples.

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