扇贝过敏原荧光免疫层析检测方法的构建与应用

目的 构建一种简便、快速、可降低非特异性荧光干扰的时间分辨免疫层析检测方法, 实现快速检测扇贝中原肌球蛋白(tropomyosin, TM)过敏原。方法 本研究采用双抗体夹心免疫层析法, 以含有铕(Eu)纳米微粒的荧光微球作为标签偶联兔抗TM多克隆抗体, 制备荧光探针并对其进行表征。以4 mg/mL兔多克隆抗体作为T线, 羊抗兔免疫球蛋白G (immunoglobulin g, IgG)作为C线组装免疫层析试纸条。结果 本研究组建的试纸条视觉检出限为0.05 μg/mL, 仪器检出限为0.01 μg/mL。试纸条除对虾蟹有交叉反应外, 对其他10余种物种无明显交叉反应。加标样品批内和批间变异系数分别为3.71%~7.94%和12.09%~12.80%, 在不含TM的4种食物基质中加入浓度由低到高的TM, 检测结果与实际加标情况相符。结论 本检测方法准确性良好、灵敏度高、特异性强, 可在多种食物基质中实现对扇贝TM的快速检测。     

量子点标记免疫层析试纸条快速检测谷物中赭曲霉毒素A

目的:根据竞争抑制免疫层析原理,构建一种基于量子点标记的免疫层析试纸条用于检测谷物中赭曲霉毒素A(Ochratoxin A,OTA)的残留量。方法:通过活化酯法将羧基功能化的量子点(Quantum dot,QD)与赭曲霉毒素A多克隆抗体(Antibody,Ab)偶联制备量子点抗体偶联物(QD-Ab);通过分别添加不同量的QD-Ab和工作液,优化量子点标记免疫层析试纸条的工作条件;通过商品化试剂盒验证该方法的有效性。结果:当QD与Ab的摩尔比为1:10时,QD-Ab荧光特性最佳;在QD-Ab和工作液添加量分别为1、10 μL时,量子点标记免疫层析试纸条结果最佳;量子点标记免疫层析试纸条的检测限为0.5 μg/L,谷物样品中的检测限为5 μg/kg,整个检测过程不超过10 min;量子点标记免疫层析试纸条特异性良好且具有有效性。结论:该方法操作简便、检测快速、结果准确灵敏,易于判断,可以满足谷物中赭曲霉毒素A残留量现场快速检测的要求。     

苯乙醇胺A荧光免疫层析试纸条的制备及特性

为通过荧光免疫层析技术制备一种快速、灵敏、特异性强的尿液中苯乙醇胺A（phenylethanolamine A,PEAA）的检测方法,采用重氮化法合成PEAA人工抗原,并通过免疫、融合、有限稀释法获得特异性抗PEAA的单克隆抗体,通过将量子点微球与PEAA偶联,制备荧光免疫层析试纸条。结果表明：PEAA免疫荧光层析试纸条的检测限为0.496 μg/L,回收率在80%～120%之间,批内、批间变异系数均小于10%,与其他类似物的交叉反应率小于5%,特异性良好。说明将荧光免疫层析技术应用到PEAA检测中是可行的,且制备的PEAA荧光免疫层析试纸条灵敏度高、准确性好、特异性好,应用前景广阔。     

呋喃西林代谢物单克隆抗体及荧光免疫层析试纸条的制备

为建立一种基于荧光免疫层析技术的呋喃西林代谢物的快速检测方法,采用活性酯法合成呋喃西林代谢物（氨基脲盐酸盐）（semicarbazide hydrochloride,SEM）人工抗原,并获得SEM的单克隆抗体,通过将量子点微球（quantumdot submicrobeads,QBs）与SEM偶联,得到QBs探针,制备荧光免疫层析试纸条。结果表明：SEM免疫荧光层析试纸条检测鱼肉组织的检测限为0.247 μg/L,不同添加质量浓度的回收率均在70%～120%之间,批内、批间变异系数均在15%以下,符合我国动物源食品中兽药残留检测方法相关指导文件的规定,且在2～8 ℃条件下可以保存6 个月以上,稳定性良好。     

荧光微球免疫层析法定量检测牛乳中酪蛋白

目的研究用荧光微球免疫层析法定量检测牛乳中的酪蛋白。方法本文以酪蛋白为抗原,免疫新西兰大白兔制备抗酪蛋白的多克隆抗体,将纯化后的多克隆抗体通过EDC介导法与荧光微球进行偶联,将滤纸、样品垫、结合垫、NC膜和吸水纸组装成试纸条,用此荧光微球免疫层析法定量检测牛乳中的酪蛋白。结果试纸条在25 min内就能判定结果,最低检测限为100 ng/mL,该方法与BSA、OVA均无交叉反应,具有很好的特异性。检测酪蛋白浓度为100.0、500.0、1000.0 ng/mL的样品,试纸条的批内回收率分别为(89.03±5.2)%、(93.47±6.9)%和(91.2±7.8)%,批间回收率分别为(87.69±6.2)%、(92.73±8.3)%、(89.82±8.5)%。结论初步建立了一种快速、方便、高灵敏度的荧光微球免疫层析方法用以检测牛乳制品中的过敏原——酪蛋白。     

量子点微球荧光免疫层析定量检测水产、畜禽肉组织中恩诺沙星

目的 建立一种快速定量检测水产、畜禽肉组织中恩诺沙星的新方法学。方法 以量子点荧光微球为新型标记探针,通过参数优化,制备获得量子点荧光微球免疫层析试纸条;进一步以T/C比值法建立定量检测恩诺沙星的新方法。结果 在最优条件下,该试纸条检测缓冲液中恩诺沙星半数抑制浓度为1.035 ng/mL,最低检测限为0.125 ng/mL;检测水产、畜禽肉组织中恩诺沙星的定量检测范围为8.75~560 μg/kg,半数抑制浓度为72.45 μg/kg,加标回收实验显示该方法定量检测组织样本中恩诺沙星的回收率在83.03~124%之间,变异系数小于15%,检测15个实际含有恩诺沙星的水产以及畜禽肉组织样本,其结果与LC/MS/MS方法检测结果高度一致;此外,加速保存实验结果显示该试纸条在室温下可保存一年。结论 本研究以量子点荧光微球为新型标记探针,构建了高灵敏定量检测恩诺沙星的新方法,实现了水产、畜禽肉组织中恩诺沙星的简单、快速、定量分析。     

基于比色和荧光双信号检测戊唑醇的免疫层析试纸条的构建

基于Au@PDA核壳纳米粒子和其对量子点荧光淬灭效应,制备了一种双信号读取模式的免疫层析试纸条,可用于定性或定量检测果蔬中戊唑醇的含量.研究了纳米金和抗体复合物的制备、试纸条传感元件的载量和缓冲液等因素对试纸条检测灵敏度的影响.在最优条件下,采用比色检测模式时,所构建双信号戊唑醇试纸条,其肉眼定性检测限为1μg/mL;...     

时间分辨荧光免疫层析试纸条在油料饼粕黄曲霉毒素B_1检测中的应用

采用黄曲霉毒素时间分辨荧光免疫层析试纸条及配套的时间分辨荧光速测仪,对油料饼粕中黄曲霉毒素B1的快速检测进行了应用研究。该时间分辨荧光免疫分析技术是基于时间分辨荧光免疫层析试纸条和载有Eu（Ⅲ）标记特异性单克隆抗体的样品瓶建立的检测技术。时间分辨荧光速测仪可内置标准曲线,直接输出检测结果。对6种油料饼粕做黄曲霉毒素B1添加回收率实验,回收率在70%~120%之间,批间、批内变异系数〈15%。在实际样品的检测中,时间分辨荧光免疫层析试纸条检测技术与液相色谱-串联质谱法相比,检测结果相对误差〈15%。时间分辨荧光免疫层析试纸条检测技术测定快速、准确,技术稳定、可靠,设备经济、小型,适用于大批量油料饼粕样品的快速检测和风险评估。     

高灵敏度磁荧光免疫层析试纸模式构建及在呕吐毒素检测中的应用

构建基于磁荧光纳米材料的免疫层析试纸模式,弥补现在免疫层析技术的不足,为更灵敏的免疫学快速检测提供技术支撑。以呕吐毒素（deoxynivalenol,DON）为靶标,采用溶剂热法制备羧基修饰的超顺磁颗粒,碳二亚胺法将磁颗粒、绿色荧光蛋白及DON单克隆抗体进行偶联,一步法制备磁荧光抗体探针,以DON人工抗原（DON-BSA）为检测线建立磁荧光免疫层析试纸。同时用胶体金标记DON单克隆抗体,以DON-BSA为检测线建立胶体金免疫层析试纸;制备的磁荧光抗体探针具有很好的磁性、荧光特性及抗体反应性,基于该探针成功制备了DON磁荧光免疫层析试纸,该试纸回归方程为y=－0.562x＋0.921,R2=0.990,IC50为5.611 ng/mL,检出限为1.089 ng/mL;制备了DON胶体金免疫层析试纸,该试纸裸眼检测灵敏度为500 ng/mL;定量检测回归方程为y=－0.543x＋1.485,R2=0.991,IC50为65.16 ng/mL,检出限为11.94 ng/mL。DON磁荧光免疫层析试纸的灵敏度是胶体金免疫层析试纸的10.96 倍。本实验建立的磁荧光免疫层析试纸模式可以同时实现样品的富集及荧光信号检测,提高检测灵敏度,并成功用于DON的检测,为磁荧光纳米颗粒广泛应用于免疫层析领域提供参考。     

大肠杆菌O157:H7量子点免疫层析试纸条的研制

研制一种大肠杆菌O157:H7量子点免疫层析试纸。利用自制水溶性量子点静电偶联大肠杆菌O157:H7单克隆抗体,将大肠杆菌O157:H7单克隆抗体和羊抗兔二抗划线于硝酸纤维素膜分别作为检测线和质控线,制备双抗体夹心法检测大肠杆菌O157:H7的量子点免疫层析试纸。该试纸条能在5min内完成检测,检测限制为1×104 CFU/mL,对常见的8种食源菌无交叉反应。基于量子点的大肠杆菌O157:H7免疫层析试纸操作简便,灵敏度和特异性较好,可用于食品快速检测。     

Reliable identification of grapevine rootstock varieties using RAPD PCR on woody samples

Since grapevine ( Vitis spp .) rootstock material is being traded increasingly as disbudded woody material a lack of distinctive morphological features on such material necessitates an alternative and reliable means of identification. Methods described here were developed for rapid and efficient extraction of DNA from woody samples rich in phenolic compounds and polysaccharides, and for subsequent identification of varieties by RAPD PCR. Using these methods, and with the application of only one selected RAPD primer, we were able to differentiate sixteen rootstock varieties, including the seven varieties most commonly used in Germany. Problems commonly encountered with reproducibility of RAPD patterns were avoided by choosing primers with a dinucleotide sequence and a high G/C content that allowed a rather high annealing temperature of 45°C. Methods described here should also be useful for other horticultural crops, especially those with woody tissues rich in phenolic compounds and polysaccharides.     

An internet compendium of analytical methods and spectroscopic information for monomers and additives used in food packaging plastics

An internet website (http://cpf.jrc.it/smt/) has been produced as a means of dissemination of methods of analysis and supporting spectroscopic information on monomers and additives used for food contact materials (principally packaging). The site which is aimed primarily at assisting food control laboratories in the European Union contains analytical information on monomers, starting substances and additives used in the manufacture of plastics materials. A searchable index is provided giving PM and CAS numbers for each of 255 substances. For each substance a data sheet gives regulatory information, chemical structures, physico-chemical information and background information on the use of the substance in particular plastics, and the food packaging applications. For monomers and starting substances (155 compounds) the infra-red and mass spectra are provided, and for additives (100 compounds); additionally proton NMR are available for about 50% of the entries. Where analytical methods have been developed for determining these substances as residual amounts in plastics or as trace amounts in food simulants these methods are also on the website. All information is provided in portable document file (PDF) format which means that high quality copies can be readily printed, using freely available Adobe Acrobat Reader software. The website will in future be maintained and up-dated by the European Commission's Joint Research Centre (JRC) as new substances are authorized for use by the European Commission (DG-ENTR formerly DGIII). Where analytical laboratories (food control or other) require reference substances these can be obtained free-ofcharge from a reference collection housed at the JRC and maintained in conjunction with this website compendium.     

Aroma characterization of various apricot varieties using headspace–solid phase microextraction combined with gas chromatography–mass spectrometry and gas chromatography–olfactometry

The characterization of the aromatic profile of several apricot cultivars with molecular tracers in order to obtain objective data concerning the aromatic quality of this fruit was undertaken using headspace–solid phase microextraction (HS–SPME). Six apricot cultivars were selected according to their organoleptic characteristics: Iranien, Orangered, Goldrich, Hargrand, Rouge du Roussillon and A4025. The aromatic intensity of these varieties measured by HS–SPME–Olfactometry were defined and classified according to the presence and the intensity of grassy, fruity and apricot like notes. In the six varieties, 23 common volatile compounds were identified by HS–SPME–GC–MS. Finally, 10 compounds, ethyl acetate, hexyl acetate, limonene, β-cyclocitral, γ-decalactone, 6-methyl-5-hepten-2-one, linalool, β-ionone, menthone and (E)-hexen-2-al were recognized by HS–SPME–GC–O as responsible of the aromatic notes involved in apricot aroma and considered as molecular tracers of apricot aromatic quality which could be utilized to discriminate apricot varieties.     

Textural Properties of Cold-set Gels Induced from Heat-denatured Whey Protein Isolates

A 9% whey protein (WP) isolate solution at pH 7.0 was heat-denatured at 80°C for 30 min. Size-exclusion HPLC showed that native WP formed soluble aggregates after heat-treatment. Additions of CaCl2 (10–40 mM), NaCl (50–400 mM) or glucono-delta-lactone (GDL, 0.4–2.0%, w/v) or hydrolysis by a protease from Bacillus licheniformis caused gelation of the denatured solution at 45°C. Textural parameters, hardness, adhesiveness, and cohesiveness of the gels so formed changed markedly with concentration of added salts or pH by added GDL. Maximum gel hardness occurred at 200 mM NaCl or pH 4.7. Increasing CaCl2 concentration continuously increased gel hardness. Generally, GDL-induced gels were harder than salt-induced gels, and much harder than the protease-induced gel.     

Tests of potential functional barriers for laminated multilayer food packages. Part I: Low molecular weight permeants

The advent of the functional barrier concept in food packaging has brought with it a requirement for fast tests of permeation through potential barrier materials. In such tests it would be convenient for both foodstuffs and materials below the functional barrier (sub-barrier materials) to be represented by standard simulants. By means of inverse gas chromatography, liquid paraffin spiked with appropriate permeants was considered as a potential simulant of sub-barrier materials based on polypropylene (PP) or similar polyolefins. Experiments were performed to characterize the kinetics of the permeation of low molecular weight model permeants (octene, toluene and isopropanol) from liquid paraffin, through a surrogate potential functional barrier (25 μm-thick oriented PP) into the food simulants olive oil and 3% (w/v) acetic acid. These permeation results were interpreted in terms of three permeation kinetic models regarding the solubility of a particular model permeant in the post-barrier medium (i.e. the food simulant). The results obtained justify the development and evaluation of liquid sub-barrier simulants that would allow flexible yet rigorous testing of new laminated multilayer packaging materials.     

Occurrence of bisphenol-F-diglycidyl ether (BFDGE) in fish canned in oil

The levels of bisphenol-F-diglycidyl ether (BFDGE) were quantified as part of a European survey on the migration of residues of epoxy resins into oil from canned fish. The contents of BFDGE in cans, lids and fish collected from all 15 Member States of the European Union and Switzerland were analysed in 382 samples. Cans and lids were separately extracted with acetonitrile. The extraction from fish was carried out with hexane followed by re-extraction with acetonitrile. The analysis was performed by reverse phase HPL C with fluorescence detection. BFDGE could be detected in 12% of the fish, 24% of the cans and 18% of the lids. Only 3% of the fish contained BFDGE in concentrations considerably above 1mg/kg. In addition to the presented data, a comparison was made with the levels of BADGE (bisphenol-A-diglycidyl ether)analysed in the same products in the context of a previous study.     

Contribution of European research to risk analysis

The European Commission's, Quality of Life Research Programme, Key Action 1—Health, Food & Nutrition is mission-oriented and aims, amongst other things, at providing a healthy, safe and high-quality food supply leading to reinforced consumer confidence in the safety of European food. Its objectives also include the enhancing of the competitiveness of the European food supply. Key Action 1 is currently supporting a number of different types of European collaborative projects in the area of risk analysis. The objectives of these projects range from the development and validation of prevention strategies including the reduction of consumers risks; development and validation of new modelling approaches; harmonization of risk assessment principles, methodologies, and terminology; standardization of methods and systems used for the safety evaluation of transgenic food; providing of tools for the evaluation of human viral contamination of shellfish and quality control; new methodologies for assessing the potential of unintended effects of genetically modified (genetically modified) foods; development of a risk assessment model for Cryptosporidium parvum related to the food and water industries; to the development of a communication platform for genetically modified organism, producers, retailers, regulatory authorities and consumer groups to improve safety assessment procedures, risk management strategies and risk communication; development and validation of new methods for safety testing of transgenic food; evaluation of the safety and efficacy of iron supplementation in pregnant women; evaluation of the potential cancer-preventing activity of pro- and pre-biotic ('synbiotic') combinations in human volunteers. An overview of these projects is presented here.     

低温带皮菜籽粕微粉的不同粒级部分的功能特性

为研究低温带皮菜籽粕微粉的不同粒级部分的功能特性,以经低温脱脂的带皮菜籽粕为原料,经微粉碎后筛分成212~425μm、150~212μm和106~150μm的3个不同粒级的微粉样品,检测这些样品的吸水性、吸油性、乳化性和乳化稳定性、蛋白质体外消化率。结果表明:1 3个不同粒级的微粉样品之间的粗纤维含量存在显著差异,表明三者的结构组成成分有一定差异。23个微粉样品的乳化活性和乳化稳定性随粒度级别的减小而显著增加(P0.01)。33个微粉样品的蛋白质体外消化率随粒度级别的减小而显著增加(P0.01)。4不同粒级带皮菜籽粕微粉样品的吸水性与吸油性受其结构组成物质不同和粒度的双重影响,与粒度的相关性不明显。     

Microbiology of food taints

Fresh and processed foods are often spoilt by the presence of undesirable flavours and odours caused by microbial action. The aim of this paper is to review the current knowledge of microbiologically induced taints that occur in a wide range of foodstuffs, including meats, poultry, fish, crustaceans, milk, dairy products, fruits, vegetables, cereals and cereal products. Examples have been chosen where the compounds responsible for the taint have been identified and sufficient data obtained to demonstrate the involvement of microorganisms. However, in some cases the full identity of the causative organism may not have been elucidated. The types of microorganisms covered by this review include bacteria, fungi, yeasts, actinomycetes and cyanobacteria. Although cyanobacteria do not in general infect foods, their presence in aqueous systems and water supplies can lead to off-flavours in aquatic organisms and processed foodstuffs. Several examples of each of these processes are discussed. Wherever possible, the likely biosynthetic pathway used by the microorganism to produce the offending compound in a foodstuff is indicated.