抗赭曲霉素A单克隆杂交瘤细胞系的建立及特性

为快速检测食物中的赭曲霉素A ,建立了抗OchratoxinA的单克隆杂交瘤细胞株。用OA -匙孔嘁血蓝素 (OA KLH)偶联物免疫 8～ 10周龄雌性Balb c小鼠后 ,取脾细胞与小鼠骨髓瘤细胞系Sp2 0融合 ,经过 3～ 4次亚克隆建立了 3个稳定分泌抗赭曲霉素A抗体的杂交瘤细胞株 ,分别命名为 10G7、10G10和 5G11。将上述 3株杂交瘤细胞分别注入Balb c小鼠腹腔 ,获得含抗赭曲霉素A单克隆抗体的腹水。将腹水用饱和硫酸铵法纯化 ,得到 10G7、10G10和 5G11单克隆抗体。其中10G10、5G11单克隆抗体的Ig亚类为IgG1,10G7单克隆抗体的Ig亚类为IgG2a。抗体腹水的稀释度为 1∶6 4× 10 6～ 1∶1 3× 10 8,参考工作浓度为 1∶1 0× 10 6～ 1∶3 2× 10 7。纯化后 10G7抗体的IgG含量为 16 4g L ,亲和常数为 9× 10 -8mol L。 10G7抗体与其它结构类似物无交叉反应 ,具有较高的特异性。     

芝麻过敏原油质蛋白O leosins单克隆抗体的制备与鉴定

利用细胞融合技术制备小鼠抗油质蛋白杂交瘤细胞,通过间接ELISA方法筛选稳定分泌特异性抗体的杂交瘤细胞株。获得10株杂交瘤细胞,分别命名为1B2、2D8、2F3、3A4、3D7、3F8、4A10、4F12、6A12、6E7,利用Ig亚类鉴定试剂盒鉴定各单克隆抗体的Ig亚型,除1B2、3D7、3F8为IgG1类外其余均为IgG2a类型。将杂交瘤细胞株注射入小鼠腹腔获取腹水,应用Protein A亲和层析法对腹水进行纯化。单抗效价有9株在2.0×105以上。ELISA和Western blotting分析表明这些单抗均能特异性识别芝麻过敏原油质蛋白。     

抗微囊藻毒素单克隆抗体的研制

为研制抗微囊藻毒素(Microcystins,简称MC)的特异性单克隆抗体,用MC-KLH偶联物免疫6～8周龄雌性BalB/c小鼠,取脾细胞与小鼠骨髓瘤细胞Sp2/0融合,经过3～4次亚克隆建立稳定分泌抗MC的杂交瘤细胞株。将杂交瘤细胞注射入BalB/c小鼠腹腔,生产出抗MC的单克隆抗体。得到了抗微囊藻毒素单克隆抗体细胞株,命名为G5,抗体亚类为IgG2a,亲和力常数2.9×10-11mol/L,分子量150KD,加标回收率在81.5%～125.0%之间,与其他结构类似物的交叉反应率<1%。筛选出的抗微囊藻毒素单克隆抗体具有较好的特异性。     

单增李斯特菌胶体金免疫层析试纸条的研制

以单增李斯特菌(LM)内化素A蛋白(InlA)单克隆抗体为基础,研制其胶体金免疫层析检测试纸条。方法 采用DNAStar软件对LM inlA全长基因编码蛋白进行抗原表位分析,截取部分inlA基因片段构建原核表达质粒,诱导表达和纯化获得重组蛋白。以该蛋白免疫BALB/c小鼠,筛选高效分泌抗InlA单克隆抗体的杂交瘤细胞株,制备单克隆抗体;以双抗体夹心的原理研制胶体金免疫层析检测试纸条,并对其特异性、敏感性、稳定性进行评价。结果 筛选到2株高效分泌抗InlA单克隆抗体的杂交瘤细胞株,抗体属于IgG1亚类,小鼠腹水抗体效价为1∶64000;研制的试纸条可与LM发生阳性反应,而与非LM李斯特菌、链球菌、鼠伤寒沙门菌、大肠杆菌O157∶H7等食源性致病菌均不发生阳性反应;LM纯培养物敏感性为2.4×10^5cfu/ml,模拟猪肉样品敏感性为4.0×10⁶ cfu/ml;4℃保存期可达16周以上。结论 研制的胶体金免疫层析试纸条具有快速、特异、敏感等优点,可以用于样品中LM的快速检测。     

抗瓜类细菌性果斑病菌单克隆抗体的制备及特性鉴定

以瓜类细菌性果斑病菌燕麦嗜酸菌西瓜亚种（Acidovorax avenae subsp. citrulli,Aac）野生型菌株SD01免疫BALB/c小鼠,间接酶联免疫吸附（enzyme-linked immunosorbent assay,ELISA）法筛选获得4 株稳定分泌抗SD01菌株的单克隆杂交瘤细胞株6F、6D、7E、4F,腹水抗体效价分别为1∶102 400、1∶102 400、1∶25 600、1∶51 200。采用饱和硫酸铵沉淀及Protein-G亲和层析法纯化腹水,十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（sodiumdodecyl sulfate-polyacrylamide gelelectrophoresis,SDS-PAGE）显示纯化后的单克隆抗体（mAb）纯度较高,纯化后单克隆抗体（2 mg/mL）效价分别为1∶12 800、1∶6 400、1∶3 200、1∶6 400,亚型鉴定结果表明4 种单克隆抗体均为IgG2 a。间接ELISA结果表明,4 种单克隆抗体对8 种果斑病菌结合能力不同：6F可以结合6 种,6D、4F可以结合8 种,7E可以结合5 种。与3 种非Aac近源种植物病原菌交叉反应情况为：6F、4F与2 种非Aac近源种植物病原菌存在交叉反应,6D、7E与3 种非Aac近源种植物病原菌均无交叉反应。     

动物源性食品中抗司帕沙星单克隆抗体的研制与鉴定

为了制备高灵敏、高特异性的司帕沙星(SPFX)单克隆抗体,鉴定免疫学特性。采用蛋白质偶联技术DCC法制备免疫抗原SPFX-BSA和包被抗原SPFX-OVA,免疫BALB/c小鼠,用间接ELISA和阻断ELISA选出产生优质多克隆抗体的的小鼠进行细胞融合,经过多次亚克隆筛选出分泌高敏感的特异性抗SPFX单克隆抗体的杂交瘤细胞株,体内诱生腹水法制备抗SPFX的单克隆抗体并对其免疫学特性进行鉴定。结果表明,间接ELISA检测显示免疫的6只小鼠效价均达1.28:10~4以上,融合后筛选出四株杂交瘤细胞株4H4-C8,4H4-C4,4H4-B1和4H4-E9;其细胞上清效价分别为1:1.4×10~3,1:8.0×10~2,1:5.0×10~2,1:6.7×10~2;4H4-C8株腹水效价为1:2.6×10~6,抗SPFX的单克隆抗体对SPFX的IC_(50)为31μg/L,且具有较好的特异性。本研究成功合成SPFX人工抗原,制备出优质的单克隆抗体,为SPFX免疫学快速检测方法的建立奠定坚实的基础。     

副溶血性弧菌单克隆抗体的制备及特性鉴定

为制备特异性强的副溶血性弧菌的单克隆抗体,解决单克隆抗体对免疫学检测产品研发的制约,以副溶血性弧菌(Vibrio parahaemolyticus)ATCC 17802标准菌株免疫Balb/c小鼠,经细胞融合、间接酶联免疫吸附测定法筛选,获得稳定分泌抗副溶血性弧菌(ATCC 17802)菌株的单克隆杂交瘤细胞株3F7D7E8C4,通过体内诱生腹水大量制备抗体,用亚类试剂盒测定抗体亚类为Ig G1;采用辛酸硫酸铵沉淀法以及亲和层析柱对腹水进行纯化,聚丙烯酰胺凝胶电泳实验鉴定单克隆抗体的纯度。制备得到腹水抗体效价为1∶16 000,纯化后抗体效价为1∶8 000,抗体敏感性IC_(50)达到10~6 CFU/m L。纯化后的抗体与12株副溶血性弧菌均能特异性结合,与其他9种非副溶血性弧菌的食源性致病菌均无交叉反应。     

玉米赤霉烯酮单克隆抗体制备及免疫分析

采用活泼酯法将玉米赤霉烯酮(Zaralenone,ZEN)分别与牛血清白蛋白(BSA)和卵清蛋白(OVA)偶联合成免疫原ZEN-BSA和包被原ZEN-OVA,经紫外扫描和聚丙烯酰胺凝胶电泳鉴定后免疫BALB/c小鼠;挑选产生高效价和高敏感性抗体的小鼠进行抗原超强免疫,取脾细胞与SP2/0细胞融合获得1株稳定分泌抗玉米赤霉烯酮单克隆抗体的杂交瘤细胞(3D8),经鉴定,该抗体亚类为IgG1,轻链为k型。经体内诱生腹水并纯化获得效价为1∶2.048×106的单克隆抗体。建立了基于单克隆抗体的ZEN间接竞争ELISA法(ic-ELISA),该方法IC50和检出限分别为22.89pg/mL和10.07pg/mL。与α-玉米赤霉烯醇、β-玉米赤霉烯醇、玉米赤霉酮、α-玉米赤霉醇和β-玉米赤霉醇的交叉反应率分别为95%、8%、12%、6%和5%,而与黄曲霉毒素B1、呕吐毒素、伏马毒素B1和赭曲霉毒素A未见有交叉反应。在玉米、大麦、小麦和燕麦中加标的回收率在86.4%~104.8%之间。该方法灵敏特异,可作为快速检测谷物中玉米赤霉烯酮的初筛方法。     

弧菌外膜蛋白OmpK单克隆抗体的制备及其特性

制备弧菌外膜蛋白K(outer membrane protein K,Omp K)单克隆抗体并对其特性进行研究。以原核表达系统表达野生株Vibrio parahaemolyticus B的Omp K免疫Balb/c小鼠,取免疫小鼠脾细胞与肿瘤细胞SP2/0进行细胞融合,采用有限稀释法和间接酶联免疫吸附测定法筛选出能够稳定分泌单克隆抗体的杂交瘤细胞株,小鼠体内诱导制备腹水,用饱和硫酸铵沉淀法和亲和层析柱纯化抗体。最终获得能稳定分泌抗Omp K的两株单克隆抗体杂交瘤细胞株Omp K-Mab-4B7、Omp K-Mab-3C5,2株杂交瘤细胞系所分泌的Mab亚类均为Ig G1,效价达1∶128 000,抗体3C5、4B7的敏感度IC50分别为2.5、5.0μg/m L。Western blotting结果显示单克隆抗体可以与本实验室12株副溶血性弧菌(V.parahaemolyticus A、B、C、D、E、F、G、H、I、J、ATCC17802、ATCC33847)的外膜蛋白有不同程度的结合反应,与4株溶藻弧菌中的3株(V.alginolyticus A、B、C)外膜蛋白能够较好的结合,与1株鳗弧菌(V.anguillarumMVM)外膜蛋白也有轻微的结合反应,实验制备的单克隆抗体可用于弧菌Omp K的基础研究和快速检测。     

酚酞单克隆抗体的制备

目的 建立免疫学快速检测减肥类保健食品中非法添加的酚酞, 制备酚酞单克隆抗体并进行评价。方法 利用碳二亚胺(carbodiimide, EDC)法合成免疫原和包被原, 用免疫原免疫Balb/C小鼠, 取小鼠脾脏与SP2/0鼠骨髓瘤细胞融合。采用竞争结合双阳性两步筛选法, 筛选出能分泌特异性抗体的杂交瘤细胞, 利用有限稀释亚克隆方法得到单株细胞; 采用体内诱生法制备腹水型单克隆抗体。利用辛酸-饱和硫酸铵法对腹水型抗体进行纯化, 利用酶联免疫吸附法鉴定纯化后的抗体。结果 成功合成了酚酞-BSA免疫原和酚酞-OVA包被原, 筛选获得酚酞杂交瘤细胞株FT/BSA/2019, 单克隆抗体的效价1×105。结论 本研究初步建立了特异性高的酚酞单克隆抗体的制备方法。     

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.     

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.     

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.     

Analysis of benzo[a]pyrene in spiked fatty foods by second derivative synchronous spectrofluorimetry after microwave-assisted treatment of samples

A simple, rapid and inexpensive method has been developed for the determination of benzo[a     

The development of reference materials for paralytic shellfish poisoning toxins in lyophilized mussel. II: Certification study

This paper describes the second part of a project undertaken to develop certified mussel reference materials for paralytic shellfish poisoning toxins. In the first part two interlaboratory studies were undertaken to investigate the performance of the analytical methodology for several PSP toxins, in particular saxitoxin and decarbamoyl-saxitoxin in lyophilized mussels, and to set criteria for the acceptance of results to be applied during the certification exercise. Fifteen laboratories participated in this certification study and were asked to measure saxitoxin and decarbamoyl-saxitoxin in rehydrated lyophilized mussel material and in a saxitoxin-enriched mussel material. The participants were allowed to use a method of their choice but with an extraction procedure to be strictly followed. The study included extra experiments to verify the detection limits for both saxitoxin and decarbamoyl-saxitoxin. Most participants (13 of 15) were able to meet all the criteria set for the certification study. Results for saxitoxin.2HCl yielded a certified mass fraction of <0.07 mg/kg in the rehydrated lyophilized mussels. Results obtained for decarbamoyl-saxitoxin.2HCl yielded a certified mass fraction of 1.59+/-0.20 mg/kg. The results for saxitoxin.2HCl in enriched blank mussel yielded a certified mass fraction of 0.48 +/- 0.06 mg/kg. These certified reference materials for paralytic shellfish poisoning toxins in lyophilized mussel material are the first available for laboratories to test their method for accuracy and performance.     

Announcing a new international peer-reviewed journal:Food Science and Human Wellness now accepting manuscript submission in English

<正>We are pleased to announce the launch of a new international peer-reviewed journal-Food Science and Human Wellness,ISSN 2213-4530,which is an open access journal,produced and hosted by Elsevier B.V.on behalf of Beijing Academy of Food Sciences.Food Science and Human Wellness is an international peer-reviewed English journal that provides a forum for the dissemination of the     

Chinese CTP Market

According to Printing and Printing Equipment Industries Association of China（PEIAC）＇s statistics to the plate manufucturer in China, in 2013, the actual offset plate production has reached 346 million square meters in China. Among them, the CTP production volume was 245 million square meters, up by 11% than that of last year; the total sales of the CTP plate was 239 million square meters, up by 13%.     

Preparatory Work of Print China 2015 is Going Smoothly

正Held at Guangdong Modern International Exhibition Center,Print China 2015 will cover 7exhibition halls,besides the original Hall No.3,4,5,6,7,the newly built F zone of Hall 3 will be used too.The total area will be140,000 square meters.Hall 3:Offset and large printing equipment,package printing equipment,post press     

INFOPRINT 2014 was Successfully Held in Dongguan

Sponsored by Printing and Printing Equipment Industries Association of China （PEIAC） and organized by Print China magazine, the Seventeenth Beijing International Printing Information Conference （INFOPRINT 2014） was successfully held on 11th Dec. 2014 at Dongguan Exhibition International Hotel.