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PubMed2004

New materials for solid-phase extraction and multiclass high-performance liquid chromatographic analysis of pesticides in grapes.

Sample preparation procedures which included the use of new aminopropyl (NH2) and octadecyl (C18) solid-phase extraction (SPE) sorbents are proposed for the simultaneous multiclass determination of the fungicide benomyl and of the herbicides tebuthiuron, diuron, simazine, atrazine, and ametryn in grapes, using single wavelength high-performance liquid chromatography. Sorbent preparation uses a fast, easy, and effective procedure to obtain silica-based materials, made by depositing polysiloxanes on a silica support followed by thermal immobilization. Recovery results of the compounds, after elution from the SPE cartridges, indicate that the most efficient system employed silica loaded with 40% of an aminofunctional polydimethylsiloxane as sorbent, using dichloromethane:methanol (95:5, v/v) as eluent. Method validation, carried out in agreement with International Conference on Harmonization directives, was performed at three fortification levels (100, 200, and 1000 microg kg(-1)). Limits of detection and quantification show that the method developed can be used to detect the pesticides at concentrations below the maximum residue levels established by Codex Alimentarius, the US Environmental Protection Agency, the European Union, and Brazilian legislation.

Chromatography, High Pressure Liquid↗
PubMed2005

High-performance liquid chromatographic determination of pesticides in tomatoes using laboratory-made NH2 and C18 solid-phase extraction materials.

A method for high-performance liquid chromatographic (HPLC) multiresidue determination of six pesticides in tomatoes was developed and validated. Silica-based C18 (octadecyl) and NH2 (aminopropyl) solid-phase extraction (SPE) sorbents, made in our laboratory, were used for sample preparation. The SPE materials were obtained by thermal immobilization of appropriate polysiloxanes onto 40 microm silica surfaces and were used in sample preparation for multiresidue analysis of the following pesticides: tebuthiuron and diuron (urea herbicides), simazine, atrazine and ametryn (triazines herbicides) and benomyl (benzimidazol fungicide). The results were compared with similar commercial materials. Reversed-phase high-performance liquid chromatography (RP-HPLC) using a Purospher RP-18 5 microm column and ACN: 0.01% aqueous NH4OH, pH 8.4 (35:65, v/v) as mobile phase, at 0.7 mlmin(-1), with 235 nm UV detection, was used for separation and quantification of the pesticides. Method validation was performed at three fortification levels (100, 200, 1000 microg l(-1)). Limits of detection and quantification show that the methods developed can be used to detect the pesticides in concentrations below the maximum residue levels (MRL) established by the Codex Alimentarius, USA, European Union and Brazilian legislations. The results showed that aminopropyl materials have a better performance than the octadecyl sorbents. Laboratory-made materials give results similar to commercial sorbents, with recoveries and precisions in agreement with directives for method validation in residue analysis.

Chromatography, High Pressure Liquid↗
PubMed2026

Proteomics as a theranostic compass in BCR::ABL1-negative myeloproliferative neoplasms: Integrating biomarker discovery with therapeutic stratification.

Classic BCR::ABL1-negative myeloproliferative neoplasms (MPNs)-polycythaemia vera, essential thrombocythaemia, and primary myelofibrosis-are clonal haematopoietic stem cell disorders with marked heterogeneity in clinical phenotype, disease trajectory, and therapeutic response. Genomic stratification by driver and cooperating mutations only partially accounts for this variability, leaving gaps in predicting thrombotic risk, fibrotic progression, leukaemic transformation, and treatment benefit. Proteomics bridges this gap by providing function-proximal readouts of protein abundance, post-translational modifications, pathway activity, and intercellular signalling that genomics and transcriptomics cannot capture, positioning it as a theranostic platform in which the same molecular readouts simultaneously inform diagnostic stratification and therapeutic decision-making. We propose a five-stage translational framework spanning from discovery-scale mass spectrometry and affinity-based plasma profiling to targeted validation, multicentre standardisation, and machine learning-integrated clinical panels. Proteomic evidence is synthesised across the following four disease axes: clonal fitness in haematopoietic stem and progenitor cells; bone marrow microenvironmental remodelling and fibrosis; chronic inflammation and thrombosis; and leukaemic transformation. We further describe how phosphoproteomics reveals resistance mechanisms to JAK inhibitors, including AXL-MAPK bypass and PP2A-autophagy-mediated tolerance, and how protein-level biomarkers (BCL2-BCL-XL, RAS-ERK, CAMK2G, and ROCK1/2) can guide individualised therapeutic selection. Affinity-based platforms (Olink PEA and SomaScan) and spatially resolved technologies (CODEX and single-cell proteomics) complement discovery proteomics. At present, however, this evidence base is constrained by small and heterogeneous cohorts, limited cross-platform reproducibility, and a scarcity of independent external validation for candidate protein panels. Realising this vision will require multicentre standardisation, analytically validated panel assays, and prospective clinical studies that translate molecular findings into decision-grade tools for patients with MPNs.

Humans↗
PubMed2006

Determination of endocrine disruptors in Kafue lechwe (Kobus leche kafuensis) samples from the Lochinvar National Park of Zambia.

Analysis of serum, whole blood and liver tissue samples from Kafue lechwe in the Lochinvar National Park of Zambia for suspected endocrine disrupting compounds revealed high concentrations for some of the compounds. 45 samples of serum, whole blood and liver tissue were extracted by liquid-liquid extraction followed by an analysis using Gas Chromatography-Electron Capture Detection (GC-ECD). The following endocrine disruptors were analyzed: deltamethrin, aldrin, endosulfan, dieldrin, pp-DDD, heptachlor, d-t-allethrin, pp-DDE, endrin and pp-DDT. For all the samples, dieldrin showed the highest concentration ranging from 1.7 to 44.4 microg/ml in serum and whole blood sample extracts and 0.10-5.1 microg/g wet weight in liver sample extracts. The most frequently detected was deltamethrin in 62% of the samples. Percent recoveries in spiked laboratory blanks ranged between 60% and 100% while calculated detection limits ranged from 0.004 to 0.21 microg/ml for all the endocrine disruptors evaluated. Where endocrine disruptors were detected, the concentrations of most of them far exceeded the maximum residue limits (MRLs) and the extraneous maximum residue limits (EMRLs) set by the Codex Alimentarius of the United Nations (UN), Food and Agricultural Organization (FAO).

Animals↗
PubMed2004

Detection and traceability of genetically modified organisms in the food production chain.

Both labelling and traceability of genetically modified organisms are current issues that are considered in trade and regulation. Currently, labelling of genetically modified foods containing detectable transgenic material is required by EU legislation. A proposed package of legislation would extend this labelling to foods without any traces of transgenics. These new legislations would also impose labelling and a traceability system based on documentation throughout the food and feed manufacture system. The regulatory issues of risk analysis and labelling are currently harmonised by Codex Alimentarius. The implementation and maintenance of the regulations necessitates sampling protocols and analytical methodologies that allow for accurate determination of the content of genetically modified organisms within a food and feed sample. Current methodologies for the analysis of genetically modified organisms are focused on either one of two targets, the transgenic DNA inserted- or the novel protein(s) expressed- in a genetically modified product. For most DNA-based detection methods, the polymerase chain reaction is employed. Items that need consideration in the use of DNA-based detection methods include the specificity, sensitivity, matrix effects, internal reference DNA, availability of external reference materials, hemizygosity versus homozygosity, extrachromosomal DNA, and international harmonisation. For most protein-based methods, enzyme-linked immunosorbent assays with antibodies binding the novel protein are employed. Consideration should be given to the selection of the antigen bound by the antibody, accuracy, validation, and matrix effects. Currently, validation of detection methods for analysis of genetically modified organisms is taking place. In addition, new methodologies are developed, including the use of microarrays, mass spectrometry, and surface plasmon resonance. Challenges for GMO detection include the detection of transgenic material in materials with varying chromosome numbers. The existing and proposed regulatory EU requirements for traceability of genetically modified products fit within a broader tendency towards traceability of foods in general and, commercially, towards products that can be distinguished from each other. Traceability systems document the history of a product and may serve the purpose of both marketing and health protection. In this framework, segregation and identity preservation systems allow for the separation of genetically modified and non-modified products from "farm to fork". Implementation of these systems comes with specific technical requirements for each particular step of the food processing chain. In addition, the feasibility of traceability systems depends on a number of factors, including unique identifiers for each genetically modified product, detection methods, permissible levels of contamination, and financial costs. In conclusion, progress has been achieved in the field of sampling, detection, and traceability of genetically modified products, while some issues remain to be solved. For success, much will depend on the threshold level for adventitious contamination set by legislation.

Animals↗
PubMed2005

Evaluation of conventionally and organically produced vegetables for high lipophilic organochlorine pesticide (OCP) residues.

The occurrence and distribution of highly hydrophobic organochlorine pesticide (OCP) in vegetables cultivated under organic and conventional conditions were evaluated. OCP residues in aerial and subterranean tissues of two varieties of lettuce and chard together with the soil where they grown were GC-ECD analyzed. SigmaDDTs>SigmaChlordane>SigmaHeptachlor>SigmaAldrins was the OCP distribution pattern in all samples. Conventional soils had higher OCP residues than organic one, even though levels were bellow 5 ng/g dry weight, indicative of low polluted agricultural environments. Vegetables accumulated OCP efficiently with residue levels 4x to 45x fold greater than those of soils. OCP tissue-dependent distribution was found to be upon the physicochemical characteristics of the pollutants (Koa and Kow). Lettuce showed a high variability in pesticide uptake regarding varieties and tillage practices. In spite of analyzed pesticides are banned or restricted, edible tissues of vegetables from both farms showed detectable residues of these compounds even though at levels below the allowed by the Codex Alimentarius. Thus, environmental conditions like presence or absence of trees, hedgerows or nearby to conventional farms influence on OCP occurrence and levels in vegetables organically grown.

Agriculture↗
PubMed2006

Incidence of aflatoxins in Iran pistachio nuts.

Aflatoxins (AF) are highly toxic and carcinogenic secondary fungal metabolites and have been detected in various food commodities including pistachio nuts. Pistachio nuts were produced in Iran during March 2002-February 2003 analyzed for aflatoxin B1 (AFB1), aflatoxin B2 (AFB2), aflatoxin G1 (AFG1) and aflatoxin G2 (AFG2) using immunoaffinity column and quantitated by HPLC and/or TLC-scanner. In this regard, 3356 pistachio nut samples were collected. After dividing samples to sub-samples, 10,068 AF analyses were done. Among 10,068 samples analyzed, AFB1 was detected in 3699 samples (36.7% of the total) with the mean and median of 5.9 (+/-41.7) ng/g and 0.1 ng/g, respectively. Total AF (AFT) was detected in 2852 samples (28.3% of the total) with the mean and median of 7.3 (+/-53.2)ng/g and 0.4 ng/g, respectively. AFB1 level in 1191 samples (11.8%) was above the maximum tolerated level (MTL) of AFB1 in pistachio nut in Iran (5 ng/g). Regarding AFT, the mean contamination level (7.3 ng/g) was lower than MTL of AFT in pistachio nut in Iran as well as lower than the proposed draft maximum level of Codex Committee on Food Additives and Contaminants for AFT (15 ng/g), and only 7.5% of samples had levels above the MTL.

Aflatoxin B1↗
PubMed2006

Management recommendations of the Federal Institute for Risk Assessment (BfR) after the international BfR symposium "Towards a Risk Analysis of Antibiotic Resistance".

The results of four expert working groups on the risk analysis process on antimicrobial resistance are described. They focus on the procedures recommended by the FAO/WHO CODEX Alimentarius Commission in 1999 including hazard identification, hazard characterization, exposure assessment and risk characterization. In all four areas, the current scientific knowledge was evaluated and recommendations for risk management options were given. They will form the basis for mitigating the health risk due to resistant microorganisms.

Animal Husbandry↗
PubMed2006

Risk analysis according to the Federal Institute for Risk Assessment international symposium "Towards a Risk Analysis of Antibiotic Resistance".

In the following, the conclusion reached by four expert working groups as regarding the aspect of risk assessment in the risk analysis process on antimicrobial resistance is outlined. Playing a central role is the procedure, which was endorsed by the FAO/WHO CODEX Alimentarius Commission in 1999, entailing hazard identification, hazard characterization, exposure assessment and risk characterization. Scientific observations in these four areas are subjected to evaluation. In essence, risk assessment decisions in this context are fundamental as they form the basis for risk management options, which in turn should reduce health risks posed by resistant microorganisms.

Agriculture↗
PubMed2006

15N spin diffusion rate in solid-state NMR of totally enriched proteins: the magic angle spinning frequency effect.

As demonstrated by means of the one-dimensional solid-state MAS exchange experiment (CODEX), the rate of the proton driven spin diffusion between backbone (15)N nuclei in totally enriched protein depends strongly on the magic angle spinning (MAS) frequency: spin diffusion at MAS frequency 16 kHz is about 4-5 times slower as compared to that at MAS frequency 1 kHz which is due to the averaging of the homo- and hetero-nuclear dipolar interactions by MAS. It is important that even at the highest MAS frequencies used in our experiments the spin diffusion rate is comparable or larger than typical values of the spin-lattice relaxation rates of backbone nitrogens in solid proteins. Thus, the precise quantitative analysis of (15)N T(1)'s in totally enriched solid proteins may lead to wrong quantitative results. On the other hand, the effectiveness of the (15)N-(15)N correlation and structure determination experiments making use of (15)N-(15)N distances can be increased by decreasing the MAS frequency as far as possible, which is counter intuitive to the commonly applied fast MAS conditions in order to reduce the dipolar-broadened line widths for increased spectral resolution.

Glycine↗
PubMed2004

Ajoene (natural garlic compound): a new anti-leukaemia agent for AML therapy.

The reputation of garlic (Allium sativum) as an effective remedy for tumours extends back to the Egyptian Codex Ebers of 1550 b.c. Several garlic compounds including allicin and its corresponding sulfide inhibit the proliferation and induce apoptosis of several human non-leukaemia malignant cells including breast, bladder, colorectal, hepatic, prostate cancer, lymphoma and skin tumour cell lines. Ajoene (4,5,9-trithiadodeca-1,6,11-triene-9-oxide) is a garlic-derived compound produced most efficiently from pure allicin and has the advantage of a greater chemical stability than allicin. Several clinical trials and in vitro studies of ajoene have demonstrated its best-known anti-thrombosis, anti-microbial and cholesterol lowering activities. Recently, topic application of ajoene has produced significant clinical response in patients with skin basal cell carcinoma. Ajoene was shown to inhibit proliferation and induce apoptosis of several human leukaemia CD34-negative cells including HL-60, U937, HEL and OCIM-1. Also, ajoene induces 30% apoptosis in myeloblasts from chronic myeloid leukaemia patient in blast crisis. More significantly, ajoene profoundly enhanced the apoptotic effect of the two chemotherapeutic drugs: cytarabine and fludarabine in human CD34-positive resistant myeloid leukaemia cells through enhancing their bcl-2 inhibitory and caspase-3 activation activities. The two key anti-leukaemia biological actions of ajoene were the inhibition of proliferation and the induction of apoptosis. Studies have shown the anti-proliferation activity of ajoene to be associated with a block in the G2/M phase of cell cycle in human myeloid leukaemia cells. The apoptosis inducing activity of ajoene is via the mitochondria-dependent caspase cascade through a significant reduction of the anti-apoptotic bcl-2 that results in release of cytochrome c and the activation of caspase-3. Since acute myeloid leukaemia (AML) is a heterogeneous malignant disease in which disease progression at the level of CD34-positive cells has a major impact on resistance to chemotherapy and relapse and the inability to undergo apoptosis is a crucial mechanism of multi-drug resistance in AML patients. The recent findings of the potent enhancing activity of ajoene on chemotherapy-induced apoptosis in CD34-positive resistant human myeloid leukaemia cells suggest a novel promising role for the treatment of refractory and/or relapsed AML patients as well as elderly AML patients. Further studies are warranted to evaluate similar enhancing effect for ajoene in blast cells from AML patients in primary cultures before its introduction in pilot clinical study.

Acute Disease↗
PubMed2005

The role of veterinary epidemiology and veterinary services in complying with the World Trade Organization SPS agreement.

The agreement on the application of sanitary and phytosanitary measures (SPS agreement) was one of the major products of the GATT's Uruguay round of multilateral trade negotiations, signed in Marrakesh on 15 April 1994. This agreement and others are part of the treaty that established the World Trade Organization (WTO). The WTO superseded the GATT as the umbrella organization for international trade (WTO, 1998a). The SPS agreement's main intent is to provide guidelines and provisions to member countries to facilitate trade while taking measures to protect human, animal or plant life or health. The agreement dictates that all sanitary measures must be scientifically based and not more restrictive than required to avoid the risk identified. The agreement recommends the use of international standards from the World Organization for Animal Health (OIE), Codex Alimentarius (CAC) and the International Plant Protection Convention (IPPC) as the basis for import requirements. If a country chooses to apply more restrictive measures than those in the international standards, it has to justify its position through a risk analysis, thus avoiding the use of sanitary and phytosanitary measures as unjustified barriers to trade. More than ever, veterinary services worldwide are faced with having to fulfill a crucial role in protecting their country's animal health status, provide sound surveillance information on the occurrence of diseases within their territories, and conduct scientifically valid risk analyses to establish justified import requirements. During the past two decades, most countries have experienced resource reduction in their veterinary services. The effect of these policies has been severe, in many cases leading to an inability of veterinary services to conduct their disease prevention and control duties. There is a clear inconsistency between the demands placed on veterinary services and the current level of funding and support they are receiving, particularly in the developing world. This paper analyzes the implications in complying with the SPS agreement and explores the role of veterinary epidemiology in developing viable alternatives that can enhance the veterinary services' ability to perform under the current economic reality. The key provisions of the SPS agreement are regionalization, risk analysis, harmonization, equivalence and transparency. The paper focuses on the contribution of epidemiology in each of these areas in the effective implementation of the SPS agreement.

Animal Diseases↗
PubMed2006

Cadmium and zinc accumulation in soybean: A threat to food safety?

A greenhouse study was conducted to quantify cadmium and zinc accumulated by soybean (Glycine max (L.) Merr.) when the metals were supplied separately and together. The highest dose of cadmium (100 mg/kg) reduced plant height and dry weight (down to 40% and 34% of control, respectively); the highest dose of zinc (2000 mg/kg) reduced plant height to 55% of control and dry weight to 70% of control. With both metals present, the plants were approximately the same size as those treated with cadmium only. The concentration of cadmium in the roots was unaffected by zinc. In other tissues, the effect of zinc on the accumulation of cadmium depended on the doses provided. At low doses, the addition of zinc reduced the concentration of cadmium in aboveground tissues to 40-50% of that found in plants exposed to cadmium only. However, when applied in high doses, the presence of zinc in cadmium-contaminated soils increased the uptake and accumulation of cadmium in aboveground tissues by up to 42%. In contrast, at high doses, the presence of cadmium in zinc-contaminated soil resulted in approximately 35% lower concentrations of zinc in all tissues. At a lower dose, cadmium had no effect on concentration of zinc in the plant tissues. The effects of high doses of one metal on the uptake of the other metal can be partially explained by the effects of one metal on the bioavailability of the other metal. In soils to which only one metal was added, bioavailable cadmium was 70-80% of the total cadmium, and bioavailable zinc was 50-70% of the total zinc. When both metals were added to the soil, 80-100% of the cadmium and 46-60% of the zinc were bioavailable. Concentrations of both metals were highest in root tissues (10-fold higher for cadmium, and up to 2-fold higher for zinc). Although relatively little cadmium was translocated to pods and seeds, the seeds of all plants (including those from control and zinc-treated plants) had concentrations of cadmium 3-4 times above the limit of 0.2 mg/kg set by the Codex Alimentarius Commission. This was surprising given that cadmium in the soil was only 1 mg/kg well below the maximum allowable amount for agricultural soil. While it is possible that more cadmium was accumulated by plants in this study than that which might occur under agricultural field conditions, these results reinforce the need to monitor concentrations of toxic metals in food crops.

Cadmium↗
PubMed2005

"Cooking the sample": radiofrequency induced heating during solid-state NMR experiments.

Dissipation of radiofrequency (RF) energy as heat during continuous wave decoupling in solid-state NMR experiment was examined outside the conventional realm of such phenomena. A significant temperature increase could occur while performing dynamic NMR measurements provided the sample contains polar molecules and the sequence calls for relatively long applications of RF power. It was shown that the methyl flip motion in dimethylsulfone (DMS) is activated by the decoupling RF energy conversion to heat during a CODEX pulse sequence. This introduced a significant bias in the correlation time-temperature dependency measurement used to obtain the activation energy of the motion. By investigating the dependency of the temperature increase in hydrated lead nitrate on experimental parameters during high-power decoupling one-pulse experiments, the mechanisms for the RF energy deposition was identified. The samples were heated due to dissipation of the energy absorbed by dielectric losses, a phenomenon commonly known as "microwave" heating. It was thus established that during solid-state NMR experiments at moderate B0 fields, RF heating could lead to the heating of samples containing polar molecules such as hydrated polymers and inorganic solids. In particular, this could result in systematic errors for slow dynamics measurements by solid-state NMR.

Journal Article↗
PubMed2006

The impact of food regulation on the food supply chain.

Food regulation in the main is aimed at protecting the consumer's health, increasing economic viability, harmonizing well-being and engendering fair trade on foods within and between nations. Consumers nowadays are faced with food or food ingredients that may derive from distant countries or continents, and with a less transparent food supply. Safety concerns must cover the range of different food chains relevant to a certain food product or product group, including all relevant producers, manufacturing sites and food service establishments within a country as well as those importing into the country. Hazard analysis at critical control points (HACCP), good manufacturing practice (GMP) and good hygiene practice (GHP) are major components of the safety management systems in the food supply chain. Principally, "a hazard" is a biological, chemical or physical agent in, or condition of, food that has the potential to cause an adverse health effect. The likelihood of occurrence and severity of the same is important for the assessment of the risk presented by the hazard to the food supply chain. The Government's regulatory mechanisms in accordance with the WTO agreements (HACCPs, sanitary and phytosanitary measures, etc.) oversee the analyses of public health problems and their association to the food supply. Under the WTO SPS Agreements and the codes of practices issued by the Codex Alimentarius Commission, there now exists a benchmark for international harmonization that guarantee the trade of safe food. Inevitably, food safety is still mainly the responsibility of the consumer.

Consumer Product Safety↗
PubMed2026

A longitudinal single-cell and spatial multiomic atlas of pediatric high-grade glioma.

Pediatric high-grade glioma (pHGG) is an incurable central nervous system malignancy that is a leading cause of pediatric cancer death. While pHGG shares many similarities with adult glioma, it comprises distinct disease entities. In this study, we longitudinally profile a molecularly diverse cohort of 16 pHGG patients through single-nucleus RNA and ATAC sequencing, whole-genome sequencing, and CODEX spatial proteomics to capture the evolution of neoplastic and microenvironmental features during disease progression and treatment. We define a set of core pHGG neoplastic cell states and observe differential tumor-myeloid interactions between malignant cell phenotypes. We find that essential neuromodulators and the interferon response are upregulated post-therapy, implicating them as malignant cell-intrinsic targets. We observe an increase in oligodendrocytes upon progression and that they coordinate spatial motifs with proneural tumor cells. This multiomic atlas of longitudinal pHGG captures features of therapy response and provides a scalable reference for the study of pediatric brain tumors.

Humans↗
PubMed2001

Wheat starch, gliadin, and the gluten-free diet.

Individuals with celiac disease generally are advised to follow a lifelong gluten-free diet and avoid consumption of the prolamins gliadin (wheat), secalin (rye), and hordein (barley). Although the designation of the diet as glutenfree may imply that the diet contains zero gluten, this is not necessarily true. In some countries (eg, United States, Canada), the gluten-free diet is completely devoid of gluten and is based on foods such as rice and corn that are naturally gluten free. In others (eg, Scandinavia, United Kingdom), the gluten-free diet may include foods such as wheat starch that have been rendered gluten free but nonetheless contain small amounts of toxic prolamins. The discrepancy in the use of foods rendered gluten free exists because the amount of toxic prolamins that individuals with celiac disease may consume without damaging the mucosa of the small intestine is unknown. Minimal research has been conducted on the toxicity of foods rendered gluten free, and there are no definitive data about whether the small amount of prolamin found in these products is safe to consume. Nonetheless, the Codex Alimentarius Standard for gluten-free foods allows a certain amount of prolamin in foods designated gluten free, and these products have been used in many countries for several decades. Well-designed, scientifically sound studies are needed to help determine the amount of toxic prolamins, if any, that may be safely consumed by individuals with celiac disease. Until this research is conducted, dietitians in the United States should continue to advise their patients against the use of wheat starch and other foods rendered gluten free.

Celiac Disease↗
PubMed1998

An innovative sandwich ELISA system based on an antibody cocktail for gluten analysis.

A cocktail sandwich ELISA based on the employ of two monoclonal antibodies (MAbs) as coating antibodies and a third MAb conjugated to horseradish peroxidase has been developed for the analysis of gluten in foods. Given that each MAb displays a wide specificity spectrum for wheat, barley, rye and oats prolamins, their combination for ELISA ensures a high crossreactivity with most of the potentially toxic gliadin, hordein, secalin and avenin protein family. One of the unprecedented features of the cocktail sandwich ELISA is that it permits for the first time analysis of barley hordeins in foods, which is unattainable using conventional or commercial ELISA kits. Besides, gliadins, hordeins and secalins are recognised to the same extent. The system provides a high detection sensitivity for gliadins, hordeins, secalins and avenins (1.5, 0.05, 0.15 and 12 ng/ml, respectively). The working linear range comprises 3-100 ng/ml with a gliadin detection limit of 1.5 ppm. This limit of detection is even better than that demanded in the latest Codex recommendation, 10 ppm. Cocktail ELISA data were contrasted with those of commercial ELISA kits and confirmed by mass spectrometry, a non-immunological technique which provides evidence for the occurrence of false positive results with the commercial kits.

Antibodies, Monoclonal↗