免疫法在食品分析中的应用

免疫法在食品分析中的应用卢大用龚镥（上海大学生命科学学院,上海２０１８００）由于当今的食品分析已包容了物理,化学,生物及微生物等诸多手段,在不同类型方法的配合及竞争中,免疫分析法可谓发展迟,后劲足,如农药等的检测中,很多美国的科技人员及企业已正式提出...     

多重PCR检测技术在食品微生物检测中的应用

现代食品行业,有很多有害的微生物严重危害食品的品质和人们的健康,甚至会引起一些严重的疾病。食品安全是对食品按其原定用途进行制作和(或)食用时不会使消费者受到伤害的一种担保。食品安全急需一些快速、敏感、特异的检测方法,以及时发现致病菌,控制污染及其可能对人体健康产生的危害。多重PCR检测技术具有快速、简便微量等优点,克服了传统检测方法操作繁琐,检测时间较长等缺点,目前正在被应用于微生物致病菌,转基因产品以及肉类品种的鉴定上,具有广阔的发展前景。本文主要是介绍了多重PCR检测技术在食品微生物检测中的原理和应用,以期望在食品微生物检测方面做出贡献。     

时间分辨荧光免疫分析在兽药残留检测中的应用

近年来,兽药残留引起食物中毒的报道日益增多,兽药残留检测的意义重大。传统的气相色谱法、液相色谱法存在前处理复杂、仪器成本昂贵等缺陷,酶联免疫吸附分析(enzyme-linked immunosorbent assay,ELISA)灵敏度也不高,而时间分辨荧光免疫分析(time-resolved fluoroimmunoassay,TRFIA)操作简便、灵敏度高,已在兽药残留检测领域引起重视。介绍了TRFIA的原理和优势,综述了其在促生长繁殖类、瘦肉增产类和杀菌驱虫类兽药残留检测中的应用,并与传统方法进行了对比,TRFIA有望取代传统的检测方法成为兽药残留检测的常规方法。     

分子生物学在食品微生物检测中的应用

食品安全是百姓关注的大问题,目前食品安全监测技术正在蓬勃发展,随着现代分子生物学技术的发展,对微生物学的研究也进入到分子、基因水平。病原微生物的检测也逐渐进入分子时代,本文介绍了应用于病原微生物检测的PCR技术、DNA指纹图谱等技术。     

实时荧光定量PCR技术在转基因食品检测领域中的应用

随着基因工程技术在农业生产中应用的深入,越来越多具有改良特征的转基因植物在全球范围内得到广泛种植,随之而来的转基因食品也迅猛发展,转基因产品大规模商业化引起了对安全性问题的担忧。为保证转基因产品标签制度的顺利实施,建立快速、准确、高通量的定量检测方法十分必要。我们综述了国内外转基因食品检测技术的研究进展,重点阐述了实时荧光定量PCR技术在转基因食品检测领域中的应用,并展望了通过构建质粒标准分子的方法来实现对更多转基因植物品系的定量检测。     

荧光定量PCR技术在临床微生物检测中的应用

荧光定量PCR技术是近些年来发展起来的一种核酸检测技术,它以灵敏度高、特异性强、重复性好、快速准确定量等特点被广泛应用于各个领域。现主要介绍荧光定量PCR技术在病毒、细菌、真菌和寄生虫4个临床微生物检测方面的应用进展。     

分子生物学方法在食品微生物检测中的应用研究

众所周知,近年来的食品安全问题层出不穷,既带来了恶劣的社会影响,更对消费者的身心健康构成了威胁,故在此背景下,食品安全问题备受关注,利用分子生物学方法检测食品微生物随之发展。对此,本文就分子生物学方法在食品微生物检测中的应用作了研究,希望对解决食品安全问题有所启示。     

基因芯片在食品检测中的应用

基因芯片技术是近十几年来生命科学领域的一大发展,其应用越来越广泛。就该技术在转基因食品、食品中的微生物、食品原料、食品中营养成分检测中的应用做一全面的回顾,因其快速、准确、高通量的特点,今后必将成为食品检测的主要方法,促进食品检测的发展,提高食品的安全性,保证人类的健康。     

荧光定量PCR技术在临床微生物检测中的应用

荧光定量PCR技术是近些年来发展起来的一种核酸检测技术,它以灵敏度高、特异性强、重复性好、快速准确定量等特点被广泛应用于各个领域。现主要介绍荧光定量PCR技术在病毒、细菌、真菌和寄生虫4个临床微生物检测方面的应用进展。     

分子生物学方法在食品微生物检测中的应用

近年来,食品安全受到广泛关注。及时、准确地检测出食品中的病原微生物是食品安全检测的重要内容,食品病原微生物的分子生物学检测技术因其特异性和灵敏性而备受瞩目。我们主要介绍了基因探针检测法、PCR检测法和基因芯片检测法的原理、开发及其在食品微生物检测中的应用。     

Soluble plasma antigen in experimental Salmonella typhimurium infection in mice

Abstract To detect and characterize Salmonella antigen in blood, outbred CF-1 female mice were inoculated intraperitoneally with S. typhimurium LT-2 and blood was assayed by ELISA for Salmonella common structural antigen. Plasma antigen was detectable early in the course of infection and increased in quantity later in the course of illness when animals showed high grade bacteremia and high counts of splenic bacteria. Antigen was associated with a cell-free plasma fraction of blood, passed through filters with cut-offs of 0.2 μ and molecular mass of 1000 kDa, and was enhanced in detectability after heating to 100°C for 15 min. Antigen was concentrated by diluting plasma 1:4 in 0.1 M EDTA, heating to 100°C, and concentrating the supernate with an ultrafiltration membrane with a molecular mass cut-off of 15 kDa. By gel filtration, antigen was associated with a peak at about molecular mass 300 kDa in heated plasma and a peak at about 380 kDa in unheated plasma. These results indicate that murine typhoid infection results in circulating soluble plasma antigen, which is heat-stable with a molecular mass of approximately 300 kDa.     

High specific activity chemiluminescent and fluorescent markers: Their potential application to high sensitivity and ‘multi-analyte’ immunoassays

The sensitivities of immunoassays relying on conventional radioisotopic labels (i.e. radioimmunoassay (RIA) and immunoradiometric assay (IRMA)) permit the measurement of analyte concentrations above ca 10⁷ molecules/ml. This limitation primarily derives, in the case of ‘competitive’ or ‘limited reagent’ assays, from the manipulation errors arising in the system combined with the physicochemical characteristics of the particular antibody used; however, in the case of ‘non-competitive’ systems, the specific activity of the label may play a more important constraining role. It is theoretically demonstrable that the development of assay techniques yielding detection limits significantly lower than 10⁷ molecules/ml depends on:

• 1 the adoption of ‘non-competitive’ assays designs;
• 2 the use of labels of higher specific activity than radioisotopes;
• 3 highly efficient discrimination between the products of the immunological reactions involved.

Chemiluminescent and fluorescent substances are capable of yielding higher specific activities than commonly used radioisotopes when used as direct reagent labels in this context, and both thus provide a basis for the development of ‘ultra-sensitive’, non-competitive, immunoassay methodologies. Enzymes catalysing chemiluminescent reactions or yielding fluorescent reaction products can likewise be used as labels yielding high effective specific activities and hence enhanced assay sensitivities. A particular advantage of fluorescent labels (albeit one not necessarily confined to them) lies in the possibility they offer of revealing immunological reactions localized in ‘microspots’ distributed on an inert solid support. This opens the way to the development of an entirely new generatio of ‘ambient analyte’ microspot immunoassays perrnitting the simultaneous measurement of tens or even hundreds of different analytes in the same small sample, using (for example) laser scanning techniques. Early experience suggests that microspot assays with sensitivities surpassing that of isotopically based methodologies can readily be developed.     

The immunoassay of gibberellins

The development of sensitive and specific solid-phase enzyme immunoassays for gibberellic acid and gibberellins A₄ and A₇ is reported. The use of antisera of high apparent affinity (K_a over 10¹⁰ l mol^-1) in conjunction with alkaline phosphatase-labeled gibberellins allows, with minimum procedural effort, the quantitative determination of sub-picogram amounts of these gibberellins. The assays reported here are applicable to most gibberellins and can be set up with 1–1.5 mg of starting material. They represent the most sensitive methods for gibberellin determination known.Abbreviations GA gibberellin - GA₃ gibberellic acid - TLC thin-layer-chromatography     

High levels of urinary midkine in various cancer patients

Midkine (MK) is a heparin-binding growth factor, which promotes growth, migration, and survival of various cells, and MK expression is increased in many human carcinomas. We determined the urinary MK level by enzyme-linked immunoassay. Taking 311pg/mg creatinine as a cut-off level, 70% of patients with various carcinomas (n=142) gave positive values, while only 5.5% of healthy volunteers (n=330) did. In case of gastric carcinoma, 17 out of 21 patients with stage 1 tumor were positive. Urinary MK levels are expected to become a convenient marker as an aid in detection of tumors.     

运用微生物法测定保健食品中的维生素B6

介绍运用微生物法测定保健食品中维生素B6的含量。GB／T5009．154—2003将微生物法规定为测定食品中维生素B6的国家标准方法Ⅲ。其原理为卡尔斯伯酵母菌需在有维生素B6存在的条件下才能生长,在一定条件下维生素B6的量与其生长量成正比关系。用分光光度仪在550nm波长下测定该菌的生长．与标准曲线相比较,从而得出该样品中维生素B6的含量。通过对国标方法作较详细的注解,并对有些地方作适当的修改,以期对需要开展此项工作的实验室及其人员会有较大的帮助。     

子宫内膜异位症患者细胞因子的表达与抗体反应

目的:探究细胞因子的表达与抗体反应在子宫内膜异位症(EMS)发生中的作用及其作为诊断指标的可能性。方法:选取子宫内膜异位症患者70例与对照组50例,检测并比较两组患者血清肿瘤坏死因子-α(TNF-α)、白细胞介素(IL-6)、单核细胞趋化因子-1(MCP-1)、血管内皮生长因子(VEGF)的表达以及抗子宫内膜抗体(EM-Ab)和转铁蛋白抗体(TRF-Ab)的阳性率。分析以上细胞因子及抗体的表达与子宫内膜异位症临床分期的关系。结果:子宫内膜异位症组TNF-α、IL-6、MCP-1与VEGF均显著高于对照组,差异具有统计学意义(P0.05);子宫内膜异位症组抗子宫内膜抗体阳性率、转铁蛋白抗体阳性率以及两种抗体均为阳性的比例均显著高于对照组,差异具有统计学意义(P0.05);TNF-α、IL-6、MCP-1与VEGF细胞因子与子宫内膜异位症分期呈正相关,分期越高、EMS病情越重则细胞因子表达水平越高(P0.05)。结论:细胞因子的表达与相关抗体反应均参与了子宫内膜异位症的发生及发展,通过检测到相关细胞因子水平的升高与自身抗体转阳可以筛选与诊断子宫内膜异位症,初步判断其临床分期和病情程度。     

时间分辨荧光免疫分析及其在临床检测中的应用

本文介绍了时间分辨荧光免疫分析法的检测原理、检测方法,分析了时间分辨荧光免疫分析仪的结构并介绍了其在临床检测方面的应用。     

应用不同方法检测食品中大肠菌群结果的比较

评价检测食品中大肠菌群不同方法。比较国家标准、行业标准和显色培养基检测方法检测大肠菌群结果的差别。国家标准和行业标准检测结果基本一致,但有差异,应用显色培养基检测大肠菌群优于目前使用的国家标准和出口食品检验行业标准方法。检测食品中大肠菌群,显色培养基检测方法快速、灵敏、特异。     

Time-resolved luminescence energy transfer immunobinding study using a ruthenium-ligand complex as a donor label

A novel immunosystem is described that exploits the effect of luminescence energy transfer from a luminescently labeled antigen to a fluorescent antibody. A luminescent ruthenium-ligand complex (D-455) with absorption/emission maxima at 456/639 nm, respectively, was employed as the donor label, and a squaraine-type cyanine label (636/655 nm), as the fluorescent acceptor label. Specifically, the system human serum albumin (HSA)/anti-HSA was studied. HSA was labeled with the donor dye D-455, and anti-HSA was labeled with the acceptor dye A-631. On formation of the antigen-antibody complex, energy transfer occurs. The radiationless energy transfer affects both the decay time of D-455 and the intensities of the emissions of both D-455 and A-631. The decay time of around 500 ns of D-455 allows frequency-domain measurements in the low kilohertz range and therefore can be based on the use of conventional optoelectronics. This also suggests gated measurements to be performed. The major difference from existing HSA immunosystems is the use of a slow decaying ruthenium-ligand complex as the donor and of a long-wave emitting cyanine acceptor dye having a high quantum yield and a decay kinetics that is governed by the rate of energy transfer from the slow decaying donor.