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1.
J Mater Chem B ; 9(44): 9153-9161, 2021 11 17.
Artigo em Inglês | MEDLINE | ID: mdl-34694310

RESUMO

Imidacloprid is the most widely used insecticide in agriculture and its intensive use over the last 30 years has caused a global concern due to its potentially toxic effects on the ecosystem. Considering the recent scientific interest in novel simple methods for imidacloprid analysis, we propose a label-free sensitive and specific localised surface plasmon resonance system for the detection of the insecticide based on 2D nanostructured metasurfaces with highly performing plasmonic properties. The specificity of the sensor proposed was achieved by covalent bio-functionalization of the metasurface using a smart and easy one-step procedure mediated by carbon disulphide. The biosensor produced was tested using a set of imidacloprid standard solutions showing a competitive limit of detection, lower than 1 ng mL-1. Our novel nanosensing configuration represents a valid and reliable solution to realize low-cost portable POC tests as an alternative to the laborious and expensive methods traditionally used for insecticide detection.


Assuntos
Inseticidas/análise , Neonicotinoides/análise , Nitrocompostos/análise , Ressonância de Plasmônio de Superfície/métodos , Anticorpos Imobilizados/imunologia , Anticorpos Monoclonais/imunologia , Ouro/química , Inseticidas/imunologia , Limite de Detecção , Nanopartículas Metálicas/química , Neonicotinoides/imunologia , Nitrocompostos/imunologia
2.
Food Chem ; 358: 129898, 2021 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-33933961

RESUMO

The sensitive detection of pesticides in complex environment is important but still challenging in presence of organic-rich water sample and food matrix. Herein, we reported a nitrile-mediated SERS immunosensor for sensitive and optical anti-interference determination of imidacloprid. Raman tag contained CN bond could provide a sharp characteristic peak in the Raman-silent spectral window (1800 ~ 2800 cm-1), which could resist the optical noises from the fingerprint region (<1800 cm-1). Aucore-Agshell bimetallic nanocuboid (AuNR@Ag) connected with antigen and Raman tag was used as Raman probe, while Fe3O4 magnetic nanoparticle functionalized with anti-imidacloprid antibody was applied as signal enhancer. Owing to the specific recognition ability between antigen and antibody, the competitive system with imidacloprid was formed. Under the optimal condition, the linear relationship was developed in the range of 10-400 nM. Finally, the SERS immunosensor was successfully applied to determine imidacloprid in real samples with recoveries from 96.8% to 100.5%.


Assuntos
Análise de Alimentos/métodos , Imunoensaio/métodos , Neonicotinoides/análise , Nitrocompostos/análise , Análise Espectral Raman/métodos , Contaminação de Alimentos/análise , Ouro/química , Imunoensaio/instrumentação , Nanopartículas Magnéticas de Óxido de Ferro/química , Neonicotinoides/imunologia , Nitrocompostos/imunologia , Praguicidas/análise , Praguicidas/imunologia , Sensibilidade e Especificidade , Análise Espectral Raman/instrumentação
3.
Talanta ; 209: 120465, 2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31892037

RESUMO

A direct competitive immunosensor for the electrochemical determination of Imidacloprid (IMD) pesticide on gold nanoparticle-modified screen-printed carbon electrodes (AuNP-SPCE) is here reported for the first time. Self-obtained specific monoclonal antibodies are immobilized on the AuNP-SPCE taking advantage of the AuNPs biofunctionalization abilities. In our biosensor design, free IMD in the sample competes with IMD conjugated with horseradish peroxidase (IMD-HRP) for the recognition by the antibodies. After that, 3,3',5,5'-Tetramethylbenzidine (TMB) is enzymatically oxidized by HRP, followed by the oxidized TMB reduction back at the surface of the SPCE. This process gives an associated catalytic current (analytical signal) that is inversely proportional to the IMD amount. The main parameters affecting the analytical signal have been optimized, reaching a good precision (repeatability with a RSD of 6%), accuracy (relative error of 6%), stability (up to one month), selectivity and an excellent limit of detection (LOD of 22 pmol L-1), below the maximum levels allowed by the legislation, with a wide response range (50-10000 pmol L-1). The detection through antibodies also allows to have an excellent selectivity against other pesticides potentially present in real samples. Low matrix effects were found when analysing IMD in tap water and watermelon samples. The electrochemical immunosensor was also validated with HPLC-MS/MS, the reference method used in official laboratories for IMD analysis, through statistical tests. Our findings make the electrochemical immunosensor as an outstanding method for the rapid and sensitive determination of IMD at the point-of-use.


Assuntos
Técnicas Biossensoriais/métodos , Imunoensaio/métodos , Nanopartículas Metálicas/química , Neonicotinoides/análise , Nitrocompostos/análise , Praguicidas/análise , Anticorpos Imobilizados/imunologia , Anticorpos Monoclonais/imunologia , Armoracia/enzimologia , Benzidinas/química , Citrullus/química , Água Potável/análise , Técnicas Eletroquímicas/instrumentação , Técnicas Eletroquímicas/métodos , Eletrodos , Contaminação de Alimentos/análise , Ouro/química , Peroxidase do Rábano Silvestre/química , Limite de Detecção , Solanum lycopersicum/química , Neonicotinoides/imunologia , Nitrocompostos/imunologia , Praguicidas/imunologia , Poluentes Químicos da Água/análise , Poluentes Químicos da Água/imunologia
4.
Allergy ; 75(2): 412-422, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31444907

RESUMO

BACKGROUND: Food allergy is associated with a high personal health and economic burden. For immunomodulation toward tolerance, food compounds could be chemically modified, for example, by posttranslational protein nitration, which also occurs via diet-derived nitrating agents in the gastrointestinal tract. OBJECTIVE: We sought to analyze the effect of pretreatment with nitrated food proteins on the immune response in a mouse food allergy model and on human monocyte-derived dendritic cells (moDCs) and PBMCs. METHODS: The model allergen ovalbumin (OVA) was nitrated in different nitration degrees, and the secondary structures of proteins were determined by circular dichroism (CD). Allergy-preventive treatment with OVA, nitrated OVA (nOVA), and maximally nitrated OVA (nOVAmax) were performed before mice were immunized with or without gastric acid-suppression medication. Antibody levels, regulatory T-cell (Treg) numbers, and cytokine levels were evaluated. Human moDCs or PBMCs were incubated with proteins and evaluated for expression of surface markers, cytokine production, and proliferation of Th2 as well as Tregs. RESULTS: In contrast to OVA and nOVA, the conformation of nOVAmax was substantially changed. nOVAmax pretreated mice had decreased IgE as well as IgG1 and IgG2a levels and Treg numbers were significantly elevated, while cytokine levels remained at baseline level. nOVAmax induced a regulatory DC phenotype evidenced by a decrease of the activation marker CD86 and an increase in IL-10 production and was associated with a higher proliferation of memory Tregs. CONCLUSION: Oral pretreatment with highly nitrated proteins induces a tolerogenic immune response in the food allergy model and in human immune cells.


Assuntos
Alérgenos/química , Alérgenos/imunologia , Hipersensibilidade Alimentar/prevenção & controle , Imunização/métodos , Nitrocompostos/imunologia , Ovalbumina/química , Ovalbumina/imunologia , Linfócitos T Reguladores/imunologia , Administração Oral , Alérgenos/administração & dosagem , Animais , Doadores de Sangue , Células Cultivadas , Citocinas/metabolismo , Células Dendríticas/imunologia , Modelos Animais de Doenças , Feminino , Hipersensibilidade Alimentar/imunologia , Humanos , Tolerância Imunológica/imunologia , Imunoglobulina E/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Nitrocompostos/administração & dosagem , Ovalbumina/administração & dosagem , Transdução de Sinais/imunologia
5.
Nutrients ; 11(10)2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31618852

RESUMO

Food proteins may get nitrated by various exogenous or endogenous mechanisms. As individuals might get recurrently exposed to nitrated proteins via daily diet, we aimed to investigate the effect of repeatedly ingested nitrated food proteins on the subsequent immune response in non-allergic and allergic mice using the milk allergen beta-lactoglobulin (BLG) as model food protein in a mouse model. Evaluating the presence of nitrated proteins in food, we could detect 3-nitrotyrosine (3-NT) in extracts of different foods and in stomach content extracts of non-allergic mice under physiological conditions. Chemically nitrated BLG (BLGn) exhibited enhanced susceptibility to degradation in simulated gastric fluid experiments compared to untreated BLG (BLGu). Gavage of BLGn to non-allergic animals increased interferon-γ and interleukin-10 release of stimulated spleen cells and led to the formation of BLG-specific serum IgA. Allergic mice receiving three oral gavages of BLGn had higher levels of mouse mast cell protease-1 (mMCP-1) compared to allergic mice receiving BLGu. Regardless of the preceding immune status, non-allergic or allergic, repeatedly ingested nitrated food proteins seem to considerably influence the subsequent immune response.


Assuntos
Alérgenos/imunologia , Lactoglobulinas/imunologia , Hipersensibilidade a Leite/imunologia , Nitrocompostos/imunologia , Animais , Linhagem Celular Tumoral , Quimases/imunologia , Quimases/metabolismo , Modelos Animais de Doenças , Feminino , Imunoglobulina A/sangue , Imunoglobulina A/imunologia , Interferon gama/imunologia , Interferon gama/metabolismo , Interleucina-10/imunologia , Interleucina-10/metabolismo , Camundongos Endogâmicos BALB C , Hipersensibilidade a Leite/sangue , Estabilidade Proteica , Proteólise , Ratos , Baço/imunologia , Baço/metabolismo , Tirosina/análogos & derivados , Tirosina/imunologia
6.
Analyst ; 144(9): 2936-2941, 2019 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-30920552

RESUMO

Imidacloprid (IMD) is one of the most used pesticides worldwide as a systemic insecticide as well as for pest control and seed treatment. The toxic and potential carcinogenic character of IMD makes its monitoring of great relevance in the field of agriculture and environment, so sensitive methodologies for in field analysis are strongly required. In this context, we have developed a competitive immunoassay for the determination of IMD using specific monoclonal antibodies followed by electrochemical detection on screen-printed carbon electrodes (SPCE). The optimized immunosensor exhibited a good reproducibility (RSD of 9%) and a logarithmic response in the range 50-10 000 pM of IMD, with an estimated detection limit (LOD) of 24 pM, which was below the maximum levels allowed by the legislation. High-Performance Liquid Chromatography-Mass Spectrometry-Mass Spectrometry (HPLC-MSMS) and Enzyme-Linked Immunosorbent Assay (ELISA) analysis were also performed for comparison purposes, where the electrochemical immunosensor exhibited a wider range of response and a lower detection limit. Matrix effects below 6.5% were obtained using tap water samples. All these characteristics make our electrochemical immunosensor a valid and advantageous tool for the in field determination of IMD.


Assuntos
Anticorpos Monoclonais/imunologia , Técnicas Eletroquímicas/métodos , Imunoensaio/métodos , Inseticidas/análise , Neonicotinoides/análise , Nitrocompostos/análise , Animais , Armoracia/enzimologia , Benzidinas/química , Técnicas Biossensoriais/instrumentação , Técnicas Biossensoriais/métodos , Carbono/química , Bovinos , Compostos Cromogênicos/química , Água Potável/análise , Técnicas Eletroquímicas/instrumentação , Eletrodos , Peroxidase do Rábano Silvestre/química , Inseticidas/imunologia , Limite de Detecção , Neonicotinoides/imunologia , Nitrocompostos/imunologia , Oxirredução , Coelhos , Reprodutibilidade dos Testes , Soroalbumina Bovina/química
7.
Free Radic Biol Med ; 129: 430-439, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30321702

RESUMO

Alzheimer's disease (AD) is a progressive form of dementia characterized by increased production of amyloid-ß plaques and hyperphosphorylated tau protein, mitochondrial dysfunction, elevated oxidative stress, reduced protein clearance, among other. Several studies showed systemic modifications of immune and inflammatory systems due, in part, to decreased levels of CD3+ lymphocytes in peripheral blood in AD. Considering that oxidative stress, both in the brain and in the periphery, can influence the activation and differentiation of T-cells, we investigated the 3-nitrotyrosine (3-NT) proteome of blood T-cells derived from AD patients compared to non-demented (ND) subjects by using a proteomic approach. 3-NT is a formal protein oxidation and index of nitrosative stress. We identified ten proteins showing increasing levels of 3-NT in CD3+ T-cells from AD patients compared with ND subjects. These proteins are involved in energy metabolism, cytoskeletal structure, intracellular signaling, protein folding and turnover, and antioxidant response and provide new insights into the molecular mechanism that impact reduced T-cell differentiation in AD. Our results highlight the role of peripheral oxidative stress in T-cells related to immune-senescence during AD pathology focusing on the specific targets of protein nitration that conceivably can be suitable to further therapies. Further, our data demonstrate common targets of protein nitration between the brain and the periphery, supporting their significance as disease biomarkers.


Assuntos
Doença de Alzheimer/diagnóstico , Linfócitos/química , Nitrocompostos/imunologia , Proteoma/imunologia , Tirosina/análogos & derivados , Idoso , Idoso de 80 Anos ou mais , Doença de Alzheimer/imunologia , Doença de Alzheimer/metabolismo , Antioxidantes/química , Antioxidantes/metabolismo , Biomarcadores/metabolismo , Complexo CD3/genética , Complexo CD3/imunologia , Estudos de Casos e Controles , Separação Celular , Proteínas do Citoesqueleto/química , Proteínas do Citoesqueleto/imunologia , Metabolismo Energético/genética , Feminino , Expressão Gênica , Humanos , Linfócitos/imunologia , Masculino , Pessoa de Meia-Idade , Nitrocompostos/química , Nitrocompostos/isolamento & purificação , Estresse Nitrosativo , Estresse Oxidativo , Cultura Primária de Células , Proteoma/genética , Proteoma/metabolismo , Transdução de Sinais , Tirosina/metabolismo
8.
Anal Chem ; 87(5): 2952-8, 2015 Mar 03.
Artigo em Inglês | MEDLINE | ID: mdl-25622025

RESUMO

The multianalyte immunoassay (MIA) has attracted increasing attention due to its high sample throughput, short assay time, low sample consumption, and reduced overall cost. However, up to now, the reported MIA methods commonly require multiple antibodies since each antibody can recognize only one antigen. Herein, a novel bispecific monoclonal antibody (BsMcAb) that could bind methyl parathion and imidacloprid simultaneously was produced by a hybrid hybridomas strategy. A chemiluminescence (CL) reaction kinetics-resolved strategy was designed for MIA of methyl parathion and imidacloprid using the BsMcAb as the unique recognition reagent. Horseradish peroxidase (HRP) and alkaline phosphatase (ALP) were adopted as the signal probes to tag the haptens of the two pesticides due to their very different CL kinetic characteristics. After competitive immunoreactions, the HRP-tagged methyl parathion hapten and the ALP-tagged imidacloprid hapten were simultaneously bound to the BsMcAb since there were two different antigen-binding sites in it. Then, two CL reactions were simultaneously triggered by adding the CL coreactants, and the signals for methyl parathion and imidacloprid detections were collected at 0.6 and 1000 s, respectively. The linear ranges for methyl parathion and imidacloprid were both 1.0-500 ng/mL, with detection limits of 0.33 ng/mL (S/N = 3). The proposed method was successfully used to detect pesticides spiked in ginseng and American ginseng with acceptable recoveries of 80-118%. This proof-of-principle work demonstrated the feasibility of MIA using only one antibody.


Assuntos
Anticorpos Biespecíficos/imunologia , Imidazóis/análise , Imunoensaio/métodos , Medições Luminescentes/métodos , Metil Paration/análise , Nitrocompostos/análise , Panax/química , Praguicidas/análise , Fosfatase Alcalina/metabolismo , Animais , Anticorpos Biespecíficos/química , Anticorpos Biespecíficos/metabolismo , Feminino , Peroxidase do Rábano Silvestre/metabolismo , Hibridomas , Imidazóis/imunologia , Imidazóis/metabolismo , Imunização , Inseticidas/análise , Inseticidas/imunologia , Inseticidas/metabolismo , Limite de Detecção , Metil Paration/imunologia , Metil Paration/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Neonicotinoides , Nitrocompostos/imunologia , Nitrocompostos/metabolismo
9.
Int Immunopharmacol ; 23(1): 283-7, 2014 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-25241247

RESUMO

Nitrated fatty acids (NFAs), thought to be produced by nonenzymatic reactions of endogenous nitric oxide (NO) with naturally present unsaturated fatty acids, have recently been identified as one of the largest single pools of biologically active NO derivatives in human plasma. As the biological role of NFAs is unknown, initial in vitro studies have shown them to be potent suppressors of inflammatory responses. The aim of the study was to collect all the literature on NFAs and its interactions with peroxisome proliferator-activated receptor gamma (PPAR-γ) and review in detail the anti-inflammatory properties of PPAR-γ interceded by NFAs. A literature survey was performed using PubMed and ScienceDirect to gather complete information on NFAs and their interactions with PPAR-γ. An exhaustive literature survey revealed that NFAs found in human plasma and urine comprises a class of cell signaling mediators that can activate PPAR-γ within its physiological concentration. NFAs exhibit anti-inflammatory and anti-fibrotic effects through PPAR-γ activation in various in vitro models tested. Besides its role in inflammation other properties of NFAs such as inhibition of enzymes, inducer of gene expression, etc., were discussed. NFAs are good electrophiles with pleiotropic biological activities. Hence NFAs can be treated as potent drug candidates.


Assuntos
Ácidos Graxos/imunologia , Inflamação/imunologia , Nitrocompostos/imunologia , PPAR gama/imunologia , Animais , Anti-Inflamatórios/uso terapêutico , Descoberta de Drogas , Ácidos Graxos/química , Ácidos Graxos/farmacologia , Humanos , Imunomodulação , Inflamação/tratamento farmacológico , Óxido Nítrico/metabolismo , Nitrocompostos/química , Nitrocompostos/farmacologia , Transdução de Sinais/imunologia
10.
Sensors (Basel) ; 12(11): 15858-72, 2012 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-23202236

RESUMO

Enzyme-linked immunosorbent assay (ELISA) based on a monoclonal antibody (MoAb) was developed for the neonicotinoid insecticide clothianidin. A new clothianidin hapten (3-[5-(3-methyl-2-nitroguanidinomethyl)-1,3-thiazol-2-ylthio] propionic acid) was synthesized and conjugated to keyhole limpet hemocyanin, and was used for monoclonal antibody preparation. The resulting MoAb CTN-16A3-13 was characterized by a direct competitive ELISA (dc-ELISA). The 50% of inhibition concentration value with clothianidin was 4.4 ng/mL, and the working range was 1.5–15 ng/mL. The antibody showed high cross-reactivity (64%) to dinotefuran among the structurally related neonicotinoid insecticides. The recovery examinations of clothianidin for cucumber, tomato and apple showed highly agreement with the spiked concentrations; the recovery rate was between 104% and 124% and the coefficient of variation value was between 1.8% and 15%. Although the recovery rate of the dc-ELISA was slightly higher than that of HPLC analysis, the difference was small enough to accept the dc-ELISA as a useful method for residue analysis of clothianidin in garden crops.


Assuntos
Anticorpos Monoclonais/imunologia , Ensaio de Imunoadsorção Enzimática/métodos , Guanidinas/análise , Inseticidas/análise , Nitrocompostos/análise , Tiazóis/análise , Cromatografia Líquida de Alta Pressão , Guanidinas/imunologia , Inseticidas/imunologia , Neonicotinoides , Nitrocompostos/imunologia , Tiazóis/imunologia
11.
J Sci Food Agric ; 92(6): 1253-60, 2012 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-22083888

RESUMO

BACKGROUND: Imidacloprid is one of the main neonicotinoid insecticides widely used in agriculture owing its broad spectrum of activity and low bioaccumulation. However, imidacloprid is toxic to honey bees and other beneficial organisms, and its residues may occur in environmental and food samples, posing a potential hazard to consumers. In this study the imidacloprid derivative bearing a three-atom length spacer was synthesized and coupled to carrier proteins. Highly sensitive and specific polyclonal antibodies against imidacloprid were successfully produced and the polyclonal antibody-based enzyme-linked immunosorbent assay (pAb-ELISA) was developed. RESULTS: The ELISA standard curve was constructed within the concentration range 0.1-100 ng mL(-1). The IC(50) value for nine standard curves was in the range 1.2-3.0 ng mL(-1) and the limit of detection was 0.03-0.16 ng mL(-1). The sensitivity of the assay was one order of magnitude higher than that in most published papers. There was almost no cross-reactivity of the antibody with four structurally related compounds (acetamiprid, nicotine, clothianidin and nitenpyram) and six other compounds, indicating that the assay displays not only high sensitivity but also high specificity. No detectable imidacloprid was found in 11 collected environmental and food samples by the assay. For imidacloprid-spiked samples, acceptable recoveries of 73.4-94.4% and intra-assay coefficients of variation of 2.2-12.8% were obtained. The assay was also validated with high-performance liquid chromatography (HPLC) and a good correlation of ELISA with HPLC was achieved. CONCLUSION: The proposed ELISA provides a sensitive, specific, simple and cost-effective quantitative/screening method for detecting imidacloprid in environmental and food samples.


Assuntos
Anticorpos , Monitoramento Ambiental/métodos , Ensaio de Imunoadsorção Enzimática/métodos , Contaminação de Alimentos/análise , Imidazóis/análise , Inseticidas/análise , Nitrocompostos/análise , Animais , Abelhas , Cromatografia Líquida de Alta Pressão , Conservação dos Recursos Naturais , Reações Cruzadas , Humanos , Imidazóis/imunologia , Concentração Inibidora 50 , Inseticidas/imunologia , Limite de Detecção , Neonicotinoides , Nitrocompostos/imunologia
12.
Sheng Wu Gong Cheng Xue Bao ; 27(6): 943-51, 2011 Jun.
Artigo em Chinês | MEDLINE | ID: mdl-22034824

RESUMO

To produce high-affinity monoclonal antibodies against pesticide imidacloprid, we synthesized the haptens 1-[(6-Carboxylethylthio-3-pyridinyl) methyl] -N-nitro-imidazolidinimine (named as H1) and 1-[(6-Chloro-3-pyridinyl) methyl]-3-carboxylpropyl-N-nitro-2-imidazolidinimine (termed as H2). And then the haptens were coupled to bovine serum albumin (BSA) and ovalbumin (OVA) for immunogen (H1-BSA) and coating antigen (H2-OVA) respectively by NHS ester method. BALB/c mice were immunized with H1-BSA conjugate. We obtained two hybridoma cell lines 2F11/A9 and 2G6/G12 secreting antibody specific for imidacloprid from the conventional hybridoma technology. The result showed that the subtypes of obtained monoclonal antibodies were IgG3 and IgG1, respectively, and the titers of ascites were up to 1:128 000. The indirect competitive ELISA indicated the IC50 values of 5.3 and 28.3 ng/mL with detection limits of 1.1 ng/mL and 7.7 ng/mL, respectively. Two monoclonal antibodies had no apparent cross reactivity with six analogous compounds. Thus, two prepared monoclonal antibodies had a very high affinity and specificity, and it could be used to develop ELISA for rapid determination of imidacloprid residue and laid a solid foundation for research and development of products for immunoassay.


Assuntos
Anticorpos Monoclonais/biossíntese , Imidazóis/imunologia , Inseticidas/imunologia , Nitrocompostos/imunologia , Animais , Especificidade de Anticorpos , Hibridomas/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Neonicotinoides
13.
Talanta ; 84(4): 1107-11, 2011 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-21530785

RESUMO

The analytical performance of a kit-based enzyme-linked immunosorbent assay (ELISA) for the determination of a neonicotinoid insecticide dinotefuran residue in rice samples is addressed. The sensitivity (I(50) value) was 5.4 ng/mL, with the limit of detection, 0.6 ng/mL and the dynamic range from 1.0 to 30 ng/mL. The ELISA showed substantially high specificity toward dinotefuran besides clothianidin (184%). For rice samples, dinotefuran was extracted with methanol and the extracts were directly determined with the ELISA because of no significant matrix interference. Good recoveries were observed and ranged from 92.5% to 113.2% with coefficients of variation below 10%. The results obtained with the ELISA correlated well with those by the HPLC method for rice samples (r>0.98). These findings strongly indicate that the evaluated and validated ELISA has a potential utility in a quick, simple, and reliable residue analysis, especially a screening method before shipment contributing to food safety.


Assuntos
Ensaio de Imunoadsorção Enzimática/métodos , Contaminação de Alimentos/análise , Guanidinas/análise , Nitrocompostos/análise , Oryza/química , Resíduos de Praguicidas/análise , Cromatografia Líquida de Alta Pressão , Reações Cruzadas , Guanidinas/imunologia , Neonicotinoides , Nitrocompostos/imunologia , Resíduos de Praguicidas/imunologia , Reprodutibilidade dos Testes , Solventes/química , Fatores de Tempo
14.
Bioconjug Chem ; 22(4): 633-41, 2011 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-21446744

RESUMO

Regulation of enzyme activity either by its substrates or by effectors is generally known as allostery. However, it has been considered hard to alter its effector specificity, despite its potential utility as a sensitive molecular sensor. To this end, we made fusion proteins consisting of an antibody variable region Fv and a circularly permutated TEM-1 ß-lactamase cpBLA. Two expression vectors encoding Fv-cpBLA with different antigen specificities were made, in which cpBLA was inserted into the linker region of the single chain Fv that specifically binds either bone-related disease marker osteocalcin (BGP) C-terminal peptide or neonicotinoid insecticide imidacloprid (ICP). The cpBLA having new termini near the active site was activated upon binding with its cognate antigen, owing to the stabilization of tethered Fv by bound antigen. As a result, both Fv-cpBLA showed specific antigen binding as well as antigen-induced enhancement in catalytic activity. Moreover, E. coli cells expressing Fv-cpBLA for ICP showed ICP concentration dependent growth in the medium containing ampicillin. The system was also applied to select for Fv-cpBLA linker mutants that confer faster growth. This will be the first of an antibody-based small molecule indicator enzyme.


Assuntos
Imidazóis/imunologia , Fragmentos de Imunoglobulinas/imunologia , Nitrocompostos/imunologia , beta-Lactamases/imunologia , Imidazóis/química , Fragmentos de Imunoglobulinas/química , Neonicotinoides , Nitrocompostos/química , beta-Lactamases/química
15.
Proc Natl Acad Sci U S A ; 107(41): 17527-32, 2010 Oct 12.
Artigo em Inglês | MEDLINE | ID: mdl-20876120

RESUMO

Neonicotinoid insecticides control crop pests based on their action as agonists at the insect nicotinic acetylcholine receptor, which accepts chloropyridinyl- and chlorothiazolyl-analogs almost equally well. In some cases, these compounds have also been reported to enhance plant vigor and (a)biotic stress tolerance, independent of their insecticidal function. However, this mode of action has not been defined. Using Arabidopsis thaliana, we show that the neonicotinoid compounds, imidacloprid (IMI) and clothianidin (CLO), via their 6-chloropyridinyl-3-carboxylic acid and 2-chlorothiazolyl-5-carboxylic acid metabolites, respectively, induce salicylic acid (SA)-associated plant responses. SA is a phytohormone best known for its role in plant defense against pathogens and as an inducer of systemic acquired resistance; however, it can also modulate abiotic stress responses. These neonicotinoids effect a similar global transcriptional response to that of SA, including genes involved in (a)biotic stress response. Furthermore, similar to SA, IMI and CLO induce systemic acquired resistance, resulting in reduced growth of a powdery mildew pathogen. The action of CLO induces the endogenous synthesis of SA via the SA biosynthetic enzyme ICS1, with ICS1 required for CLO-induced accumulation of SA, expression of the SA marker PR1, and fully enhanced resistance to powdery mildew. In contrast, the action of IMI does not induce endogenous synthesis of SA. Instead, IMI is further bioactivated to 6-chloro-2-hydroxypyridinyl-3-carboxylic acid, which is shown here to be a potent inducer of PR1 and inhibitor of SA-sensitive enzymes. Thus, via different mechanisms, these chloropyridinyl- and chlorothiazolyl-neonicotinoids induce SA responses associated with enhanced stress tolerance.


Assuntos
Arabidopsis/imunologia , Ascomicetos/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Guanidinas/farmacologia , Imidazóis/farmacologia , Inseticidas/farmacologia , Nitrocompostos/farmacologia , Ácido Salicílico/imunologia , Tiazóis/farmacologia , Arabidopsis/microbiologia , Cromatografia Líquida de Alta Pressão , Regulação da Expressão Gênica de Plantas/imunologia , Guanidinas/química , Guanidinas/imunologia , Imidazóis/química , Imidazóis/imunologia , Inseticidas/química , Inseticidas/imunologia , Espectrometria de Massas , Estrutura Molecular , Neonicotinoides , Nitrocompostos/química , Nitrocompostos/imunologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Ácido Salicílico/metabolismo , Tiazóis/química , Tiazóis/imunologia
16.
Free Radic Biol Med ; 46(8): 989-1003, 2009 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-19200454

RESUMO

The signaling mediators nitric oxide ( NO) and oxidized lipids, once viewed to transduce metabolic and inflammatory information via discrete and independent pathways, are now appreciated as interdependent regulators of immune response and metabolic homeostasis. The interactions between these two classes of mediators result in reciprocal control of mediator synthesis that is strongly influenced by the local chemical environment. The relationship between the two pathways extends beyond coregulation of NO and eicosanoid formation to converge via the nitration of unsaturated fatty acids to yield nitro derivatives (NO(2)-FA). These pluripotent signaling molecules are generated in vivo as an adaptive response to oxidative inflammatory conditions and manifest predominantly anti-inflammatory signaling reactions. These actions of NO(2)-FA are diverse, with these species serving as a potential chemical reserve of NO, reacting with cellular nucleophiles to posttranslationally modify protein structure, function, and localization. In this regard these species act as potent endogenous ligands for peroxisome proliferator-activated receptor gamma. Functional consequences of these signaling mechanisms have been shown in multiple model systems, including the inhibition of platelet and neutrophil functions, induction of heme oxygenase-1, inhibition of LPS-induced cytokine release in monocytes, increased insulin sensitivity and glucose uptake in adipocytes, and relaxation of preconstricted rat aortic segments. These observations have propelled further in vitro and in vivo studies of mechanisms of NO(2)-FA signaling and metabolism, highlighting the therapeutic potential of this class of molecules as anti-inflammatory drug candidates.


Assuntos
Ácidos Graxos/metabolismo , Inflamação , Óxido Nítrico/metabolismo , Nitrocompostos/metabolismo , Transdução de Sinais/imunologia , Animais , Citocinas/metabolismo , Ácidos Graxos/imunologia , Regulação da Expressão Gênica , Heme Oxigenase-1/genética , Heme Oxigenase-1/imunologia , Heme Oxigenase-1/metabolismo , Humanos , Óxido Nítrico/imunologia , Nitrocompostos/imunologia , Estresse Oxidativo , PPAR gama/imunologia , PPAR gama/metabolismo , Processamento de Proteína Pós-Traducional/imunologia , Ratos , Vasodilatação
17.
Immunopharmacol Immunotoxicol ; 28(3): 491-500, 2006.
Artigo em Inglês | MEDLINE | ID: mdl-16997797

RESUMO

A nasal formulation of mometasone furoate (MF) is advantageous in avoiding systemic activity characteristic of glucocorticoids when it is applied topically. To confirm antiallergic effects of this glucocorticoid formulation elaborately, we investigated whether the drug can suppress the production of IgE antibodies and related cytokines. It we showed that IgE production induced in mice immunized via intranasal route was significantly reduced when the mice were administered MF intranasally. Further, MF was effective in inhibiting production of type-2 helper T cell cytokines in vivo and in vitro. These results provide a immunopharmacological basis for clinical efficacy of this drug.


Assuntos
Formação de Anticorpos/efeitos dos fármacos , Imunoglobulina E/imunologia , Interleucina-4/biossíntese , Pregnadienodiois/farmacologia , Administração Intranasal , Animais , Antialérgicos/administração & dosagem , Antialérgicos/imunologia , Antialérgicos/farmacologia , Formação de Anticorpos/imunologia , Linhagem Celular , Hipersensibilidade a Drogas/imunologia , Hipersensibilidade a Drogas/metabolismo , Hipersensibilidade a Drogas/prevenção & controle , Imunização/métodos , Imunoglobulina E/metabolismo , Imunoglobulina G/imunologia , Imunoglobulina G/metabolismo , Injeções Subcutâneas , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Furoato de Mometasona , Nitrocompostos/administração & dosagem , Nitrocompostos/imunologia , Ovalbumina/administração & dosagem , Ovalbumina/imunologia , Pregnadienodiois/administração & dosagem , Pregnadienodiois/imunologia , Células Th2/efeitos dos fármacos , Células Th2/imunologia , Células Th2/metabolismo
18.
J Immunol Methods ; 310(1-2): 159-70, 2006 Mar 20.
Artigo em Inglês | MEDLINE | ID: mdl-16499922

RESUMO

The emission of soot during combustion processes used in transportation, manufacturing, and power generation is of increasing concern because of its serious adverse health effects. In particular, the ability to modulate the immune system has recently been established. In the present investigation, an artificial soot sample that was prepared by fragmentation of acetylene in a laser-induced plasma was used as an antigen for the immunization of a rabbit. A highly sensitive competition curve in an indirect competitive ELISA using a benzo[a]pyrene-BSA conjugate as a coating antigen could be constructed for benzo[a]pyrene with an IC50 of 2.94 mug/l (11.65 nmol/l). In contrast to the high affinity, the soot antiserum dilution (antibody titer) of 1:750 was rather low. The cross-reactivity was tested with 16 parent polycyclic aromatic hydrocarbons, 7 nitrated polycyclic aromatics, and 3 methylated, hydroxylated or butyric acid derivatives. The results obtained suggest that the vertebrate immune system can respond to an immunization with soot by the generation of high affinity IgG class antibodies against polycyclic aromatics. It is likely that antibodies are raised against the molecular structures which form the framework of the soot particles and not against adsorbed and extractable polyaromatic compounds. The experiments suggest that if soot is considered a T-independent antigen, the isotype switch, essentially from IgM to IgG, could have been caused by co-inoculation with a T-dependent antigen, i.e., mycobacteria contained in Freunds complete adjuvant. However, at the cellular level the mechanism remains to be uncovered.


Assuntos
Carbono/imunologia , Imunoglobulina G/imunologia , Imunoglobulina M/imunologia , Nitrocompostos/imunologia , Hidrocarbonetos Policíclicos Aromáticos/imunologia , Acetileno/química , Animais , Reações Cruzadas , Ensaio de Imunoadsorção Enzimática , Imunoglobulina G/biossíntese , Imunoglobulina G/sangue , Imunoglobulina M/biossíntese , Imunoglobulina M/sangue , Concentração Inibidora 50 , Masculino , Microscopia Eletrônica de Transmissão , Coelhos
19.
J Agric Food Chem ; 51(7): 1823-30, 2003 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-12643637

RESUMO

An enzyme-linked immunosorbent assay (ELISA) was developed for the neonicotinoid insecticide thiamethoxam, 3-(2-chlorothiazol-5-ylmethyl)-5-methyl-4-nitroimino-1,3,5-oxadiazinane. Three antisera were raised from rabbits immunized with the hapten-KLH conjugate. On the basis of the computational analysis of hapten candidates, the hapten with a spacer arm on the thiazolyl ring of thiamethoxam was synthesized to elicit thiamethoxam-specific antisera. The hapten was 3-[2-(2-carboxyethylthio)-5-ylmethyl]-5-methyl-4-nitroimino-1,3,5-oxadiazinane. Antisera were characterized with indirect competitive ELISA. Cross-reactivity and effects of organic solvents, pH, and ionic strengths were evaluated. The antiserum was specific for thiamethoxam and tolerant of up to 5% acetonitrile and 5% acetone. Various ionic strengths and pH values in the tested ranges had negligible effect on the assay performance. Under the optimized conditions, the half-maximal inhibition concentration (IC(50)) and the limit of detection were approximately 9.0 and 0.1 microg/L of thiamethoxam, respectively. ELISA analysis of stream and tap water samples showed an excellent correlation with the fortification levels.


Assuntos
Ensaio de Imunoadsorção Enzimática/métodos , Inseticidas/análise , Nitrocompostos/análise , Oxazinas/análise , Animais , Especificidade de Anticorpos , Haptenos/química , Haptenos/imunologia , Hemocianinas/imunologia , Soros Imunes/imunologia , Modelos Moleculares , Estrutura Molecular , Neonicotinoides , Nitrocompostos/química , Nitrocompostos/imunologia , Oxazinas/química , Oxazinas/imunologia , Coelhos , Tiametoxam , Tiazóis , Poluentes da Água/análise
20.
Zh Mikrobiol Epidemiol Immunobiol ; (5): 15-20, 1984 May.
Artigo em Russo | MEDLINE | ID: mdl-6540021

RESUMO

The immunogenic properties of the dihapten and monohapten derivatives of polyethylene glycol with different nitroaromatic groupings were studied. 2,4-dinitrophenyl, 2, 4, 6-trinitrophenyl and trinitrophenyl-ethyl groupings were used as hapten groups. The injection of monohapten compounds was found to induce the accumulation of antibody-forming cells secreting antibodies to trinitrophenyl in the spleen of normal and athymic nude mice. As early as on day 3 the number of antibody-forming cells considerably exceeded their background level, the process of B-cell activation being, to a certain extent, thymus-independent. Dihapten compounds were not immunologically active. The effect rendered by the nitroaromatic derivatives of polyethylene glycol, revealed in this study, is linked with the known capacity of polyethylene glycol to adsorb on the surface of cell membranes.


Assuntos
Antígenos/imunologia , Haptenos/imunologia , Nitrocompostos/imunologia , Nitrobenzenos/imunologia , Polietilenoglicóis/imunologia , Animais , Células Produtoras de Anticorpos/efeitos dos fármacos , Antígenos T-Independentes/imunologia , Dinitrobenzenos/imunologia , Relação Dose-Resposta Imunológica , Eritrócitos/efeitos dos fármacos , Eritrócitos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Camundongos Nus , Peso Molecular , Trinitrobenzenos/imunologia
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