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1.
用增殖与浓缩后的牛病毒性腹泻病毒(BVDV)分别免疫健康牛犊和家兔,制备牛抗BVDV和兔抗BVDV超免疫血清,并用层析法纯化牛抗BVDV IgG和兔抗BVDV IgG,建立了检测BVDV的双抗体夹心ELISA快速检测方法.结果表明:该检测方法的最佳反应条件为:牛抗BVDV IgG包被浓度为100μg/mL,兔抗BVDV IgG最佳工作浓度为50μg/mL,样品反应时间为90min,酶标抗体工作浓度为1∶8 000,以D490nm≥0.177作为阳性判定标准.该ELISA检测方法的重复性CV小于10%,最低检测限为1.87μg/mL,与牛轮状病毒、牛冠状病毒、猪瘟病毒、犊牛大肠杆菌和沙门菌等病原无交叉反应,该ELISA试剂在室温下至少可保存6个月.用该ELISA方法和IDEXX公司抗原检测试剂盒对甘肃省不同地区牛场156份临床粪便样品进行了检测,阳性检出率分别为17.3%和16.7%.表明本试验建立的ELISA方法特异性强、敏感性高且重复性好,可用于BVDV的快速检测.  相似文献   

2.
The allelopathic potential of Chinese pine (Pinus tabulaeformis Carr.) against its own seed germination and seedling growth was tested with aqueous extracts (0.01, 0.02, 0.05, and 0.10 g·mL−1) obtained from different organs (roots and litter needles) at different individual ages (12, 52, and 110 years old). The results showed that root and litter extracts had different effects on seed germination and seedling growth, and the effects varied with the concentrations, the organs, and the tree age of extracts. The strongest stimulatory effect on seed germination of Chinese pine was exposed to 0.02 g·mL−1 root extract from the 110 years old Chinese pine trees and exposed to 0.02 g·mL−1 litter extract from the 12 years old Chinese pine trees. Meanwhile, the strongest stimulatory effect on growth of Chinese pine seedlings was exposed not only to 0.01 g·mL−1 root extracts from the 110 years old Chinese pine but also to 0.01 g·mL−1 litter extract from the 12 years old Chinese pine. The promoting effect of the extracts of root on seed germination and seedling growth increased in the order of 12, 52, and 110 years old. The promoting effect of the extracts of litter on seed germination and seedling growth increased in the order of 110, 52, and 12 years old. Our results suggested that litter leachates or root exudates of Chinese pine may influence the natural regeneration within Chinese pine stands via the release of allelochemicals into the environment.  相似文献   

3.
为筛选牛病毒性腹泻-黏膜病病毒(Bovine viral diarrhea virus,BVDV)gP48蛋白的单链抗体,本研究利用噬菌体展示技术构建了gP48蛋白单链抗体克隆文库,通过微孔筛选法对抗体库进行3轮富集淘筛,利用ELISA技术测定单链抗体的亲和力,对亲和力较高的克隆进行基因测序分析和结合活性测定,旨在获得gP48蛋白的单链抗体。结果表明:1)成功构建了库容量为4.3×107的噬菌体单链抗体库,其重组率为83.3%,经三轮筛选对噬菌体抗体库进行富集,富集倍数为1.43×104;2)优化了间接ELISA方法,筛选到3株与BVDV gP48蛋白具有高亲和力的单链抗体以及其基因序列,通过IgBLAST分析显示,3株单链抗体均为鼠源IgG。综上,从gP48蛋白的单链抗体库中筛选获得3个具有高亲和力的单链抗体,可用于后续BVDV检测方法的建立。  相似文献   

4.
The effects of prey density (0.1, 0.3, 1, 3 and 10 prey·mL−1) on the growth and survival of hybrid snakehead (Channa argus × C. maculate) larvae were investigated. The larvae were divided into three groups with different body lengths of 0.68 cm, 1.50 cm and 3.20 cm, respectively. The growth of the hybrid snakehead larvae in all three groups increased with prey density increasing from 0.1 to 1 prey·mL−1. The specific growth rate (SGR) was the highest when the prey density was 1 prey·mL−1. When prey density was higher than 1 prey·mL−1, SGR of larvae in Group I (the larvae of early development stage) decreased, while no significant change was observed in those of Group II and Group III. The survival rates of hybrid snakehead larvae in all three groups were high (91.11%–100%) and not significantly affected by the prey densities except in Group I with the highest prey density (10.0 prey·mL−1) which was significantly lower than the others. Body size was not sensitive to prey density. The optimum prey density was confirmed at 1 prey·mL−1 in all the treatments. __________ Translated from Journal of Huazhong Agricultural University, 2007, 26(3): 367–370 [译自: 华中农业大学学报]  相似文献   

5.
The wheat × maize system is one of the most effective ways to produce haploids in wheat. Whether and how it could be successfully applied in practical breeding mostly depends upon the efficiency of haploid embryo production. To perfect the protocols of haploid embryo induction, the efficiency of haploid embryo production between in vitro culture of cut plant and intact plant growth for hybrid spikes with two F1 wheat hybrids and two maize varieties was compared. Effects of different cutting plant times and formulas of nutrient solutions for cut plant culture on haploid embryo formation were also studied. Results indicated that the embryo rate of in vitro culture was 3.29 times that of intact plant growth, with the figures of 31.6% vs 9.6%, respectively. The optimal time for cut plant culture was 24 h after pollination. Formulas of nutrient solutions significantly affected the efficiency of haploid embryo induction. With an embryo rate of 0–35.5%, adding calcium phosphate in the culture solution at 3 g·L−1 could raise the caryopsis and embryo rates. According to this study, the best medium for cut plant culture was: 100 mg·L−1 2,4-D+ 40 g·L−1 sucrose + 10 mg·L−1 silver nitrate + 8 mL·L−1 sulfurous acid + 3 g·L−1 calcium phosphate, with which a caryopsis rate of 95% and an embryo rate of about 30% could be obtained. __________ Translated from Journal of Triticeae Crops, 2008, 28(1): 1–5 [译自: 麦类作物学报]  相似文献   

6.
A method is described for the determination of ultratrace cadmium by coupling a continuous flow vapor generation system with in situ preconcentration technique and electrothermal atomic absorption spectrometry (ETAAS). A graphite tube coated with Ir as permanent chemical modifier was used for trapping cadmium vapor species. The effects of the flow rates of carrier gas and sample injection in vapor generation systems on the trapping measurement for cadmium were respectively investigated. Graphite tubes with different characteristic surfaces were comparatively studied for trapping cadmium vapor. The experimental results showed that the permanent chemical modifier of Ir is an alternative to the thermolabile modifier of Pd for simplifying the trapping measurement. The trapping efficiency of cadmium on the graphite tube coated with Ir was estimated. The trapping temperature and time were also investigated. A detection limit (3σ) of 0.005 μg·L−1 was obtained for this proposed method. The relative standard deviation (RSD) was 1.4% for 0.5 μ·L−1 of Cd (n = 11). This method can be applied to the determination of ultratrace cadmium in food and environmental samples with good agreement between the certified and found values.  相似文献   

7.
The herbicidal mechanism of the components extracted from Pythium aphanidermatum was examined in this study. Component I was isolated using the HPD500 macroporous adsorption resin and HPLC. Its impact on seed germination and plant growth of weeds was determined and the contents of MDA, superoxide anion radical, and the activities of hills and roots were examined. The root length of weed plants was inhibited under illumination while the stem height was inhibited evidently under darkness. The relative electric conductivity of Digitaria sanguinalis and Amaranthus retroflexus under illumination was 94.55 μS·cm−1 and 58.75 μS·cm−1, respectively, whereas that under darkness was 85.25 μS·cm−1 and 36.25 μS·cm−1, respectively. The MDA contents of Digitaria sanguinalis and Chlorella pyrenoidosa were 0.08385 μmol·L−1 and 0.1742 μmol·L−1 under illumination, respectively, while those were 0.0129 μmol·L−1 and 0.01935 μmol·L−1 under darkness, respectively. Simultaneously, superoxide anion radical content was higher under illumination than under darkness. These results showed the photosynthesis was affected by component I extracted from Pythium aphanidermatum.  相似文献   

8.
抗牛病毒性腹泻病毒单克隆抗体的研制   总被引：1，自引：0，他引：1  
将含有牛病毒性腹泻病毒1型(BVDV1)囊膜糖蛋白E2编码基因的真核表达质粒pEC143免疫BALB/c小鼠,应用淋巴细胞杂交瘤技术,以重组E2蛋白纯化产物为检测抗原,经间接ELISA法筛选和3次有限稀释法克隆,得到1株能稳定分泌抗BVDV1和BVDV2单克隆抗体的杂交瘤细胞株.该3D8株单抗能识别BVDV1标准毒株(NADL、OregonC24V)、BVDV2标准毒株(890、104)、2株BVDV1分离株和3株BVDV2分离株,而不能识别牛轮状病毒、牛疱疹病毒1型,显示出良好的特异性.阳性杂交瘤细胞的上清液、腹水ELISA抗体效价分别为2~9和10×2~7,为IgM亚类.其单克隆抗体杂交瘤细胞株的获得,为建立单抗介导的检测BVDV1和BVDV2抗原和抗体的血清学方法和实现标准化诊断奠定基础.  相似文献   

9.
牛布鲁氏菌间接ELISA抗体检测方法的建立   总被引：2，自引：0，他引：2  
【目的】建立高通量牛布鲁氏菌间接ELISA诊断方法,为科学高效诊断牛布鲁氏菌病提供技术手段。【方法】以提纯的猪布鲁氏菌S2株脂多糖(LPS)作为包被抗原,采用棋盘滴定法确定了抗原包被浓度、包被液、抗原包被时间、封闭液、样品稀释液、血清稀释倍数、兔抗牛酶标抗体稀释液、兔抗牛酶标抗体稀释浓度、最佳TMB底物反应时间等参数,初步建立了牛布鲁氏菌间接ELSIA方法。并用上述条件初步建立的方法对176份牛布鲁氏菌病阳性血清和132份牛布鲁氏菌病阴性血清进行检测。检测结果经SPSS17.0软件分析,构建受试者工作特征曲线(ROC)及曲线下面积,确定敏感性和特异性;通过Youden指数确定阴阳性判定的临界点。使用质控阴、阳性血清评价试剂盒的批内和批间重复性。用本研究建立的ELISA试剂盒及商品化试剂盒同时对临床上1 200份牛血清样本进行比对检测,比较其符合率。【结果】通过棋盘法确定的试剂盒中各组分的最佳条件为:抗原包被浓度为10μg·m L-1,最适包被液为碳酸盐缓冲液,最佳包被时间为2—8℃、16 h,最适血清稀释度为1﹕50,最佳封闭液为含有3%明胶的PBST,最佳样品稀释液为含有0.5%蔗糖的PBST,最佳兔抗牛酶标抗体稀释液为含有5%马血清的PBST,最佳兔抗牛酶标抗体工作浓度为1﹕20 000,最佳TMB底物反应时间为15 min。按上述条件建立的间接ELISA方法检测176份牛布鲁氏菌病阳性血清和132份牛布鲁氏菌阴性血清,并统计结果后使用SPSS17.0软件分析该方法受试者工作特征曲线(ROC)线下面积为0.98。最佳判定临界点为样本OD值/阳性OD值(S/P)×100%=20%。在此临界值上时,敏感性为97.7%,特异性为95.5%。对1 200份临床样本的比对检测显示,本研究建立的间接ELISA方法与进口商品化试剂盒的总符合率为96.25%。【结论】建立了特异性和敏感性良好的牛布鲁氏菌间接ELISA抗体检测方法。  相似文献   

10.
本研究首次成功地建立了检测PPV抗原的间接斑点酶联免疫吸附试验(Dot—ELISA)标准化程序,在所确定的最适条件下,对纯化PPV抗原的最低检出量为2.5ng/dot,其敏感性为血凝试验(HA)的125倍,但稍低于银加强胶体金检测法(SECGA)。PPV阳性猪血清的特异性阻断试验及与猪瘟病毒,猪伪狂犬病病毒,猪巴氏杆菌的交叉反应试验证明,该方法对PPV抗原的检测具有特异性。以该方法检测PPV人工感染兔样本和自然感染猪样本,PPV抗原的阳性检出率分别为肾样本100％(17/17)、81.82％(9/11),肝样本100％(16/16)、56.52％(13/23)。20份随机样本的间接Dot—ELISA检测结果与病毒分离和鉴定结果相符。 试验证明间接Dot—ELISA对PPV感染的检测具有简单、快速、经济、敏感性高、特异性强、重复性好和便于推广应用等优点,适用于大批量抗原样本的检测,是PPV感染快速诊断和流行病学调查的有效方法。  相似文献   

11.
In a completely randomized block design experiment, 16 ruminally cannulated male sheep with body weights of (40 ± 2.1) kg were fed twice daily (8:00 and 16:00) with concentrate and forage (50:50 on dry matter (DM) basis). Dietary treatments were supplemented with intraruminal doses of powdered Yucca schidigera extract (YSE) at the levels of 0 (control), 100, 200 and 300 mg · kg−1. On days of 15, 16 and 17 after feeding, ruminal content was sampled at 0, 2, 4, 6 and 8 h after dosing (8:00), and blood samples were collected at the end of experiment (the days 18 and 19 after feeding). Results showed that the treatment groups’ acidity was not affected (P = 0.13) by YSE. Comparing to the control, the ruminal propionate concentration was increased by YSE addition in a dose-dependent manner by up to 29.8% (P < 0.05), and the acetic concentration was decreased by up to 17.5% (P < 0.05). The ruminal ammonia concentration 2 hours after feeding was higher (P < 0.05) in sheep fed without YSE (increased by 17.57 mg· 100 mL−1) than those fed with YSE at 200 mg · kg−1 (6.77 mg · 100 mL−1 increase in NH3) and at 300 mg · kg−1 (6.50 mg· 100 mL−1 increase in NH3). Protozoal populations in the rumen were lower (P < 0.05) with the YSE feeding dose at 300 mg · kg−1 than the control. The serum chemistries were not different among treatments (P > 0.05) and were within the normal physiological ranges for sheep 19 days after feeding. The study indicated that 200 mg· kg−1 and 300 mg· kg−1 YSE groups had particular suppressing effects on ruminal ammonia concentration, ammonia-N concentrations and protozoal populations. The effect of YSE on ruminal fermentation could be attributed to the selective inhibitory effect on rumen microbial species. High level (300 mg · kg−1) YSE as feed additives resulted no negative impact on sheep in our tests.  相似文献   

12.
This paper describes the chemical composition and antimicrobial activity of the essential oil of Syzygium aromaticum against Vibrio spp. (n = 6), Edwardsiella spp. (n = 21), Aeromonas spp. (n = 2), Escherichia coli (n = 2), Flavobacterium spp. (n = 1), Salmonella spp. (n = 2), Streptococcus spp. (n = 1) and Pseudomonas spp. (n = 1) isolated from aquaculture sites as well as seven reference strains of bacteria, namely, Escherichia coli (ATCC 25922), Citrobacter freundii (ATCC 8090), Aeromonas hydrophila (ATCC 49140), Pseudomonas aeruginosa (ATCC 35032), Streptococcus agalactiae (ATCC13813), Edwardsiella tarda (ATCC 15947) and Yersinia enterocolitica (ATCC 23715). Nowadays, most antibiotics are no longer effective in controlling diseases in aquaculture, especially fish systemic bacterial diseases, due to increasing incidences of antibiotic resistance among pathogenic bacteria. Furthermore, many countries have banned antibiotics in aquaculture use due to public health concerns and environmental hazards. Therefore, this study was carried out to evaluate the potential of the essential oil of S. aromaticum as an alternate commercial antibiotic to antimicrobial agents against fish systemic bacteria in aquaculture. The essential oil of S. aromaticum was prepared using a steam distillation method, and the chemical composition was analysed using Gas chromatography-mass spectroscopy (GC-MS). Minimum inhibitory concentration (MIC) values of the essential oils against the tested bacteria were determined using the broth two fold micro dilution method, with kanamycin and eugenol as positive controls. The MIC values of the essential oil of S. aromaticum ranged from 0.015 μg·mL−1 to 0.062 μg·mL−1 against the tested bacterial isolates. A total of nine chemical compounds were detected in the essential oil, with eugenol (49.0%) and caryophyllene (7.5%) being the major compounds. The results of the present study indicate that the essential oil of S. aromaticum shows a huge potential to substitute commercial antibiotics as antimicrobial agents for aquaculture use.  相似文献   

13.
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14.
采用特异性好的鼠源抗猪流行性腹泻病毒(PEDV)单克隆抗体为捕获抗体,兔源多克隆抗体为检测抗体,建立PEDV双抗体夹心ELISA检测方法。结果显示,该方法的最佳反应条件为:抗PEDV单克隆抗体E1包被质量浓度4.40μg·mL~(-1),37℃包被2h,采用5%BSA封闭液封闭1h,兔抗PEDV抗体工作质量浓度为5.91μg·mL~(-1),酶标二抗稀释度为1∶2000,以OD450nm≥0.381作为阳性判定标准。该ELISA方法对猪轮状病毒和猪传染性胃肠炎病毒无交叉反应。敏感度可达30μg·mL~(-1)(5×103.12);重复性变异系数小于10%。采用该方法和RT-PCR方法同时检测临床样品42份,阳性样品符合率为92.30%,表明建立的PEDV双抗体夹心ELISA检测方法具有特异性好、敏感性高和方便快捷等优点,可用于PEDV快速检测。  相似文献   

15.
本文介绍了应用酶标单抗(HRP-McAb)的涂片法、双抗体夹心 ELISA、斑点 ELISA 和微量细胞培养联合免疫酶测定法检测猪瘟病毒抗原,结果4种方法检出率基本一致,具有快速、特异、敏感、准确、简便、经济和判断客观的诊断价值,可作为疫区流行病检查和检测猪瘟病毒抗原应用。  相似文献   

16.
In order to develop a new method for determining the phosphorus (P) digestibility in vitro in feedstuffs by dialysis tube, a L32(49) orthogonal experiment with eight factors (4 levels for each factor) and a single factor experiment on the enzymatic reactivity were carried out. The sequence of significance of the eight factors on sample-P dialyzability was as follows: trypsin digestion for 6 h, dialyzing solution at 100 mL, pH of pepsin solution at 2.5, pepsin concentration at 2000 U·mL−1 pepsin digestion for 100 min, at temperature of 35°C, trypsin concentration at 1625 U·mL−1, and pH of trypsin solution at 6.5, respectively. And in vitro dialyzabilities of P in soybean meal, barley, sorghum, peanut meal, and rapeseed meal were (36.91 ± 0.58)%, (27.28 ± 0.94)%, (26.95 ± 0.58)%, (30.51 ± 0.83)%, and (20.82 ± 1.09)%, respectively. __________ Translated from Journal of South China Agricultural University, 2007, 28(3): 85–89 [译自: 华南农业大学学报]  相似文献   

17.
Glandless upland cotton has an important economic value. Embryogenic calli and regenerated plants were obtained from the hypocotyl explants of glandless upland cotton seedlings, cultivar Jisheng1. The results indicated that somatic embryogenesis was significantly influenced by the types of auxin and cytokinin. 2, 4-D was advantageous to induce cotton callus, but embryogenic callus could not be obtained on the 2, 4-D medium. Embryogenic calli were also not obtained on the MSB sold medium with the combination of IBA and BA. However, embryogenic calli were induced when the hypocotyl explants were cultured on the IBA and KT medium. More than 31% of the hypocotyl segments produced embryogenic calli when the MSB medium was supplemented with 1.0 mg·L−1 IBA and 0.5 mg·L−1 KT. Embryogenic calli with somatic embryos could be observed within three months. Somatic embryo germination and maturation occurred on the hormone-free MSB medium with 1.0 g·L−1 Gln and 0.5 g·L−1 Asn. A number of regenerated plants could be obtained in six months. In the present study, a simple and efficient system was established to induce a number of embryogenic calli and regenerate plantlets from hypocotyl explants.  相似文献   

18.
[目的]为牛病毒性腹泻病毒抗体快速检测提供敏感、特异检测方法.[方法]重组质粒pET-(1-345)转化宿主菌BL21(DE),以IPTG进行诱导表达,使外源基因获得了较高水平的表达; Westem一blot检测表达产物反应原性,同时进行特异性检测;将收集的纯培养物进行超声波裂解后以His bind kit蛋白质纯化试剂盒纯化回收目的蛋白,再以纯化后的目的蛋白作为包被抗原进行方阵滴定试验,确定目的蛋白作为包被抗原的最佳稀释度.[结果]重组蛋白表达可达菌体蛋白总量的19.7;,且具有较好的反应原性和特异性,以此方法建立的间接ELISA方法对收集的约43头份血清样品进行检测, 检测阳性结果与进口试剂盒检测结果符合率达到93.55;.[结论]以BVDV E2重组蛋白作为抗原建立的间接ELISA检测方法是可行的,为其进一步的标准化莫定基础.  相似文献   

19.
To identify the allelopathic effect of Flaveria bidentis (L.) Kuntze (F. bidenti) on other plants, the effects of different extracts from F. bidentis on the growth of several plants were studied by bioassay. Results showed that the water extracts inhibited the growth of corn (Zea mays L.), wheat (Triticum aestivum L.), cotton (Gassypium Hirsutum L.), soybean (Glycine hispida L.), peanut (Arachi shypogaea L.), rice (Oryza sativa L.), crabgrass (Digitaria sanguinalis L.) and rigweed (Amaranthus retroflexus L.), with the most reactive indexes found in root and stem of cotton at −0.85 and −0.88, respectively, at a concentration of 0.2 g·mL−1. However, the water extracts accelerated the growth of rice. In addition, the reactive indexes of the extracts of petroleum ether chloroform, ethyl acetate, acet and alcohol were higher than that of the water extracts, and that of the acet extracts was the highest. The melting point of the refined acetone extract ranged from 192.5°C to 193.5°C, and its maximum absorbing wavelength was 220 nm. This extract was found to be herbicide-active and played an inhibitory role in the growth of crabgrass and rigweed at concentrations of 1000, 500, 100 and 50 mg·L−1. __________ Translated from Journal of Agricultural University of Hebei, 2007, 30(6): 63–67 [译自 : 河北农业大学学报]  相似文献   

20.
An efficient extraction method by ultrasound-assisted extraction (UAE) was developed for the extraction of polychlorinated biphenyls (PCB 28, 52, 101, 118, 130, 153, 180) from various shellfish. Analytes were determined by gas chromatography-mass spectrometry in selected ion monitoring (SIM) mode. All the samples were extracted by n-hexane and decontaminated by concentrated sulfuric acid. The resulting samples were determined by quantitative detection by comparing with external standards. The average recoveries of seven kinds of PCBs ranged from 80.92% to 93.89%, and the lowest detectable limit was 0.08-0.03 μg·kg−1. The average concentrations of total PCBs were 14.12–30.61 μg·kg−1 in the samples. This method was highly effective in reducing the cost and time for the pretreatment of samples. This method may be applied for the screening and monitoring of shellfish for organic pollutants in coastal waters.  相似文献   

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