抗黄曲霉毒素单链抗体基因的克隆与表达

为降低黄曲霉毒素抗体制备成本,在已有的能分泌抗黄曲霉毒素单克隆抗体的杂交瘤细胞系8F6的基础上,成功克隆得到了该单克隆抗体的重链（VH）和轻链可变区（VL）基因片段。通过重叠延伸PCR的方法将轻、重链可变区基因连接,并引入连接肽（Linker）编码序列,构建VH-Linker-VL结构的单链抗体（ScFv）基因,并将该基因克隆到噬菌体表达载体pCANTAB 5E上,使单链抗体以噬菌体展示形式在大肠杆菌TG1中表达。间接竞争ELISA方法检测到该ScFv对黄曲霉毒素B1的抑制率（IC50）值为0.57ng/mL,表明该单链抗体与亲本鼠单抗有相同的抗原结合特异性,且具有很高灵敏度。     

黄曲霉毒素分析方法研究进展

黄曲霉毒素分析方法研究进展江苏省微生物研究所（２１４０６）宓晓黎，成恒嵩自从１９世纪６０年代，英国Ｎｏｒｆｏｒｋ发生大批火鸡中毒事件，在几个月之内死亡１０万多只；非洲的肯尼亚和乌干达也在小鸭中发生类似的疾病，死亡很多。最初人们怀疑这种疾病是田细菌和病...     

抗黄曲霉毒素B_1单抗独特型抗体的制备

目的研制黄曲霉毒素B1标准品的替代品并应用于酶联免疫吸附试验(ELISA)。方法在研制了抗黄曲霉毒素B1单抗的基础上,将抗体用无花果蛋白酶进行酶切,制备出抗黄曲霉毒素B1单抗F(ab’)2片段,并以F(ab’)2片段作为免疫原免疫日本大耳白兔,制备出抗黄曲霉毒素B1单抗独特型抗体,并对该独特型抗体进行鉴定。结果该独特型抗体是Ab2β型,与黄曲霉毒素B1存在内影像关系。结论该抗体可以替代黄曲霉毒素B1标准品应用于ELISA检测。     

黄曲霉毒素脱除方法研究进展

黄曲霉毒素污染是影响花生、玉米、小麦等农作物安全性的重要原因之一。根据黄曲霉毒素的理化特性可用物理、化学或者生物方法进行脱除。本文对黄曲霉毒素的主要脱除方法进行了综述, 以期为保障农作物及其制品的安全性提供参考。     

高特异性黄曲霉毒素B_1单克隆抗体的制备及特性研究

本研究将黄曲霉毒素B1转化为半缩醛B2a,在硼氢化钠（NaBH4）还原作用下与载体蛋白偶联制备完全抗原。将制备的完全抗原免疫Balb/c小鼠,经4次免疫后取其脾脏与小鼠骨髓瘤细胞Sp2/0细胞融合,采用半固体培养基筛选后鉴定,获得杂交瘤细胞株3A12,抗体的灵敏度可达6.1±0.025ng/mL,抗体与其它黄曲霉毒素B2、G1及G2的交叉反应率依次为7.8%、20.2%及0.6%,与黄曲霉毒素M1交叉反应率小于0.1%。本研究为研发花生等农产品黄曲霉毒素B1特异性免疫分析技术及产品奠定了重要基础。     

黄曲霉毒素脱毒研究进展

黄曲霉毒素是危害食品安全和人体健康最主要的真菌毒素之一,全世界每年因黄曲霉毒素造成的粮食及其他农产品损失严重。此外,在我国,黄曲霉毒素还是影响储藏中药材安全标准的重要污染物。由于黄曲霉毒素的危害性极高,关于黄曲霉毒素的污染防治是保证食品药品安全的研究重点之一。本研究综述了近年来关于食品中黄曲霉毒素脱毒与降解研究的最新进展以及各种方法涉及可能的降解产物和降解机制,涵盖了以物理,化学和生物3大类脱毒技术的分析和总结,除传统技术手段之外,还对于采用天然产物防治黄曲霉毒素、不同脱毒手段协同作用的相关研究进行了展望,以期为黄曲霉毒素防治和脱毒相关研究工作提供借鉴,促进后续研究的开展。     

黄曲霉毒素生物防治的研究进展

黄曲霉毒素是由真菌在适宜的条件下产生的有毒次级代谢物,是毒性最强的生物毒素之一,它能够广泛污染粮食、饲料、食品,使其产量降低与产品品质受到影响,严重地威胁人类与动物的生命安全.黄曲霉毒素的防治一直是各国研究热点.从微生物的抑制作用,如:微生物的拮抗作用、微生物的吸附降解作用;以及添加活性物质,如:添加微生物提取物、添加酶制剂等方面综述了黄曲霉毒素的生物防治研究现状.     

粮油食品中黄曲霉毒素去除方法研究进展

黄曲霉毒素广泛存在于粮油食品中,给全球农作物生产造成巨大经济损失。黄曲霉毒素具有极强的毒性和致癌性,严重威胁着人类的健康,迫切需要寻找一种有效、安全、环保控制黄曲霉生长或降解黄曲霉毒素的方法。对物理方法、化学方法和生物方法抑制黄曲霉生长或降解黄曲霉毒素的研究进行了综述。     

黄曲霉毒素去毒方法研究进展

黄曲霉毒素毒性极强,具有致癌、致畸和致突变作用,在食物链内污染范围广,严重威胁到人类的健康和饮食安全.目前对黄曲霉毒素去毒方法研究较多,其中安全有效的生物学去毒方法具有很好的发展前景.发展安全、高效且环保的黄曲霉毒素去毒方法是今后研究的热点.     

生物法防治黄曲霉毒素B1研究进展

黄曲霉毒素具有极强的致癌、致畸和致突变作用。受黄曲霉毒素污染的粮食不仅给人体和动物的健康带来严重危害,也给食品、畜牧等行业造成巨大经济损失。目前,黄曲霉毒素降解方法研究已成为各国科研工作者的一个热点。生物法降解黄曲霉毒素较物理、化学等方法具有安全系数高、特异性强、绿色清洁等独特优势,被视为最具发展前景和潜力的降解方法。利用微生物菌体的吸附作用、微生物酶解作用、微生物代谢作用等对黄曲霉毒素进行脱除,已逐渐成为生物法消减黄曲霉毒素的重要研究方向。通过筛选和改造获得可脱除黄曲霉毒素的优良菌株;通过分离、提取、纯化等手段获得高纯度和高活性的微生物源解毒产物;通过改进和优化获得最佳脱除工艺等,已成为研究生物法脱除黄曲霉毒素的重要手段和突破口。本文主要对国内外生物法防治黄曲霉毒素B_1的研究进行综述,并对今后生物防治的发展方向提出一些设想。     

钙制剂研究进展

钙是人体内重要的生命元素,直接影响人的身体健康。本文综述了钙的主要生理功能、吸收机制以及影响钙吸收的因素,主要介绍了目前钙制剂的研究情况及其评价。     

Indirect determination of aflatoxin B1 in beer via a multi-commuted optical sensor

This paper reports the determination of aflatoxin B₁ (AFB₁), one of the most carcinogenic substances known. A multi-commuted flow injection–solid phase spectroscopy (FI–SPS) system combined with photochemically induced fluorescence (PIF) was developed, for the first time, for its quantitative determination. A strongly fluorescent degradation product was obtained on-line by irradiation with ultraviolet light. The determination was carried out by measuring the fluorescence intensity of the photo-product at 353/424 (λ _ex/λ _em), once retained on C₁₈ silica-gel filling the flow-cell. A linear dynamic range of 0.09–12?µg?l^?1, detection limit as sensitive as 29?ng?l^?1 and a relative standard deviation (RSD) of 1.4% were obtained. The method proposed was satisfactorily applied to the determination of AFB₁ in different types of beer (normal and non-alcoholic). Hydrophobic compounds were eliminated from beer samples and AFB₁ was extracted with acetonitrile by solid-phase extraction on C₁₈ sorbent. Recoveries of the target compound from spiked beers were between 94 and 106%. The results obtained in the analysis of real samples are in good agreement with those provided by a reference chromatographic method.     

Exposure to aflatoxins in Japan: risk assessment for aflatoxin B1

The intake of total aflatoxins (AFT) and aflatoxin B₁ (AFB₁) from food in Japan was estimated from AFT and AFB₁ concentration and frequency data in 24 foods (884 samples) from a 3-year retail market survey from the summer of 2004 to the winter of 2006, and by food consumption data from the National Health and Nutrition Survey performed in 2005. The AFT and AFB₁ survey revealed that peanut, peanut products, cocoa, chocolate, pistachio, white pepper, red pepper, almond, job's tears, buckwheat and corn grits are considered to be contributors of AFT (or AFB₁) intake in Japan (maximum AFB₁ (AFT) levels ranged from 0.21 to 28.0 µg kg^?1 (from 0.21 to 9.0 µg kg^?1)) in AFT-contaminated food. A probabilistic approach using the Monte Carlo method was carried out to simulate an estimate of the AFT (or AFB₁) intake distributions in each age group in Japan. In this study, AFB₁ intake ranged from 0.003 to 0.004 ng kg^?1 body weight day^?1 (from lower to upper limits), and the potential risk for cancer using a formula devised by the Joint Food and Agricultural Organization/World Health Organization (FAO/WHO) Expert Committee on Food Additives (JECFA) was estimated at 0.00004–0.00005 person/year/100,000 persons, even though this was in the higher levels (95.0th percentile) of the consumer population. The results suggest that the current dietary intake of AFB₁ in Japan has no appreciable effect on health.     

小麦球蛋白单克隆和多克隆抗体制备及建立酶联免疫吸附法快速检测技术

目的建立小麦球蛋白的ELISA检测技术,用于蛋白质掺假以及过敏原成分的快速检测。方法从麦胚粉中提取球蛋白,抗原免疫Balb/c小鼠进行4次免疫后,通过脾脏细胞杂交瘤技术及间接ELISA筛选制备单克隆抗体,同时制备兔抗小麦球蛋白的多克隆抗体。通过棋盘滴定法,初步确定单克隆抗体和多克隆抗体的最佳工作浓度,建立双抗夹心ELISA。结果通过免疫和杂交瘤技术获得了抗小麦球蛋白的单克隆抗体,纯化后抗体的效价均达到1:107,通过免疫兔制备的多克隆抗体经纯化后效价在1:2.4×105左右,所建立的双抗体夹心ELISA方法最低检测限为10 ng/m L,与其他物种的蛋白不发生交叉反应。结论本文建立的双抗体夹心ELISA方法具有良好的特异性和灵敏度,为建立乳品中小麦成分掺假及过敏原成分快速检测提供理论依据。     

应用抗黄曲霉毒素单链抗体检测酱油中黄曲霉毒素B1

利用本实验室制备的抗黄曲霉毒素B1的单链抗体(ScFv),通过棋盘实验确定了抗原抗体的最适工作浓度,在此之上根据间接竞争酶联免疫法(ELISA)绘制标准曲线,检测酱油中AFB1的含量;通过改变样品的盐浓度及pH来确定其对ELISA检测结果的影响。研究结果表明,利用ScFv检测黄曲霉毒素的最小检测值为0.10ng/mL,平均加标回收率在84%～109%之间,本文建立的ELISA方法在pH5～8,盐浓度小于10%时较稳定。本文建立的利用抗AFB1的ScFv检测黄曲霉毒素含量的方法方便快捷,稳定性较好,并且成本较低,适合于食品中黄曲霉毒素的检测。     

基于适配体生物传感器检测黄曲霉毒素B1的研究进展

随着生活水平的提高, 人们对生命健康格外注重, 而由生物毒素引起的食物中毒的悲剧却频繁发生, 因此得到了社会的广泛关注。简便快速、特异性高、易携式的生物毒素检测仪器成为研究人员研究的热点。适配体(aptamer)作为一种新型识别分子, 具有亲和力高、稳定性强、制备成本低、特异性强等明显优势, 已成功应用于食品中有害物质的识别检测。随着色谱技术、免疫化学以及光、电等领域的不断发展, 将适配体与靶物质的特异性识别结合过程转换为易于检测的物理化学信号, 能够实现灵敏度高、微型化、高通量和现场检测的快速分析。本文主要从试纸条法、荧光、电化学传感器等方面以黄曲霉毒素B1(aflatoxin B1, AFB1)为靶标的适配体在食品安全检测分析中的应用进行了综述, 以期为进一步研究提供参考。     

番茄红素制剂化技术研究进展

番茄红素抗氧化活性强,具有延缓衰老、抗动脉粥样硬化、防癌、抗癌、预防心血管疾病等效果。但番茄红素稳定性差、脂溶性的特点导致其生物利用度低,限制了其应用领域。因此,通过制剂化处理提高番茄红素的稳定性、水溶性、生物利用度具有重要价值。综述了番茄红素制剂化技术,主要包括微胶囊化技术、脂质体技术、纳米分散体技术、包埋技术、乳化技术和超临界流体技术。     

Effect of the inclusion of adsorbents on aflatoxin B1 quantification in animal feedstuffs

The extraction efficiency of aflatoxin B₁ (AFB₁) in cattle feed containing nine adsorbents (ADSs) was investigated using two organic/aqueous solvents composed of methanol/water (80/20 v/v; MeOH) and acetone/water (85/15 v/v; AC). Samples were obtained including a highly AFB₁-contaminated (HC) and a low-level AFB₁-contaminated (LC) feedstuff (15.33 and 7.57 µg kg^–1, respectively), nine ADSs (four clay minerals; one yeast cell wall-based product; one activated carbon and three commercial ADS products) at two different levels of inclusion (10 and 20 g kg^?1). After solvent extraction and immunoaffinity column clean-up, all samples were analysed for AFB₁ by high-performance liquid chromatography (HPLC) with fluorescence detection. For each contamination level (HC and LC), the data obtained were analysed using a factorial arrangement in a completely randomized design. Means were compared with the correspondent controls using the Dunnett's test. No statistical difference was found in AFB₁ levels of feedstuffs not containing ADSs when extracted with AC or MeOH, even if numerically higher values were obtained with AC. A dose-dependent effect (p < 0.01) of ADSs inclusion was observed on AFB₁ recoveries that were lower when the higher ADS level (20 g kg^?1) was included in the HC and LC feedstuffs. Higher AFB₁ recoveries were obtained using AC compared with MeOH, both in HC (75.0% versus 12.0%, respectively) and in LC (84.0% versus 22.8%, respectively) ADSs containing feedstuffs. However, when the activated carbon and the sodium bentonite were included in feeds, lower AFB₁ concentrations with respect to control values (p < 0.001 and <0.05, respectively) were obtained also using AC. The data obtained in this study indicate that routine use of the MeOH solvent for AFB₁ analysis of unknown feedstuffs, can produce misleading results if they contain an ADS.