分子印迹固相萃取和丝网印刷电极联用技术检测猪尿中盐酸克伦特罗

实验采用本体热聚合的方式合成盐酸克伦特罗的分子印迹物(molecularly imprintedpolymers,MIPs),通过扫描电镜、吸附实验和分子印迹固相萃取研究该印迹材料的性能。用盐酸克伦特罗分子印迹材料作为固相萃取剂,优化固相萃取条件,采用差分脉冲伏安法(DPV)对洗脱液进行电化学性能测试。结果表明,此印迹物的平衡吸附常数(b)为5.14mL/mol,电化学传感器的电流值与盐酸克伦特罗的浓度在0.056～2.78mg/L浓度范围内呈现良好的线性关系,检测下限为0.005mg/L,盐酸克伦特罗在猪尿样品中的回收率为89.3%～99.6%。此萃取方法也呈现出良好的选择性和稳定性。     

大型溞胆碱酯酶间接非竞争酶联免疫吸附定量分析法的建立

用达到电泳纯的大型溞(Daphniamagna)胆碱酯酶(cholinesterase,ChE)免疫BALB/c小鼠,得到效价达1∶8000的鼠抗大型溞ChE多克隆抗血清;通过优化抗原抗体反应条件,建立定量分析大型溞ChE含量的间接非竞争酶联免疫吸附法(indirectandnon‐competitiveenzyme‐linkedimmunosorbentassay).结果表明:此方法检测灵敏度为24ng爛mL-1;经免疫交叉反应鉴定,所获抗血清与5、10μg爛mL-1标准蛋白质和其他生物如花翅摇蚊、尖额溞、斑马鱼(脑、脑除外的整体)、家蚕、意大利蜜蜂、赤子爱胜蚯蚓、非洲爪蟾(蝌蚪)等来源的ChE交叉反应率分别为:<0畅25%、<0畅25%、 2畅13%、 10畅19%、3畅88%、 3畅56%、 2畅81%、 3畅93%、 5畅00%和2畅75%,具较高的特异性.说明用本试验得到的抗血清建立的间接非竞争酶联免疫吸附法可用于大型溞体内ChE含量的测定,以利于在农药暴露环境下大型溞体内ChE活性的精确检测.     

基于电化学传感器检测农副产品黄曲霉毒素B1的研究进展

黄曲霉毒素B1是农副产品中最常见的真菌毒素之一,具有诱变、致畸、免疫抑制和致癌的作用。为了减少黄曲霉毒素B1带来的危害,开发可靠的方法来检测农副产品中的黄曲霉毒素B1十分重要。传统的检测方法难以满足现代农业中现场、快速检测的需求,而电化学传感器具有制备简单、便于携带、灵敏度高和选择性强等特点,因而备受关注。根据识别元件的不同,黄曲霉毒素B1的电化学传感器可以分为基于适配体的传感器、基于免疫反应的传感器、基于分子印迹的传感器。研究者们又根据不同的感知策略、纳米材料研发了多种类型的黄曲霉毒素B1电化学传感器。该研究综述了近5年来（2019—2023年间）黄曲霉毒素B1电化学传感器的研究进展,列举了具有代表性的实例,讨论了基于不同识别元件的传感器的传感机理和不同的信号产生策略,对这些传感器的性能进行了对比。并分析了不同识别元件的优缺点,探讨了基于不同原理的黄曲霉毒素B1电化学传感器的不足之处和发展方向,如需要开发新型的识别元件、研发新的纳米材料并提高纳米材料的制备技术、不同传感策略的联合应用、研发多种毒素同时检测的传感器、简化修饰步骤提高传感器稳定性等,以期为实现黄曲霉毒素B1电化学传感器的实际应用提供参考。     

Longevity of Mycobacterium bovis in brushtail possum (Trichosurus vulpecula) carcasses,and contact rates between possums and carcasses

Abstract

AIM: To determine, for a variety of environmental conditions, how long Mycobacterium bovis might remain viable inside the carcass of a brushtail possum (Trichosurus vulpecula) that died of bovine tuberculosis (Tb), and to measure the rate of contact between free-ranging possums and possum carcasses.

METHODS: Lesions of M. bovis were simulated by inoculating excised spleens weighing 0.5–1 g with 0.2 mL liquid culture containing approximately 5 x 10⁷ cfu M. bovis/mL. Simulated lesions were inserted into possum carcasses (n=48) at the peripheral lymph nodes. Carcasses were placed in the field at two sites (a tussock grassland and a podocarp-broadleaved forest site) and in two seasons (summer and winter) for up to 62 days. Survival rates of M. bovis were estimated by sampling the simulated lesions over time, and culturing the recovered lesion to determine if any viable M. bovis bacteria were present.

The time taken for a free-ranging possum to first encounter a dead possum in its home range was estimated by live-trapping possums and fitting them with proximity loggers (n=13). A ‘contact’ was recorded if these possums came within 40–50 cm of proximity loggers fitted to possum carcasses.

RESULTS: There were strong seasonal and site effects in the survival rate of M. bovis in possum carcasses. In the grassland habitat, no viable bacilli were cultured from any carcass after 3 days in summer, whereas in winter all samples were culture-positive for the first 20 days, and some were still positive after 27 days. The survival rates for forest habitat were intermediate between the results for grassland, and there were no culture-positive carcasses after 9 days in summer or 27 days in winter.

In summer, infected carcasses (n=6) were first encountered by possums a mean 1.9 (range 0.4–6.7) days after placement.

CONCLUSIONS: Possum carcasses were contacted by free-ranging possums within the period that viable M. bovis were shown to survive in a carcass. The risk of such infection is likely to be most significant in winter or in areas with microhabitats where the survival of M. bovis is high. However, the generally low survival rate of M. bovis in possum carcasses and the low frequency of possum-to-carcass contacts indicate this route of transmission alone could not maintain Tb in a possum population.     

副猪嗜血杆菌间接ELISA抗体检测试剂盒的研制与应用

【背景】副猪嗜血杆菌（Haemophilus parasuis,HPS）是猪的上呼吸道病原菌,引起格拉瑟氏病,主要引起断奶前后和保育阶段的猪发病,通常见于5—8周龄的青年猪,发病率一般为10%—15%。该菌有15种血清型,目前主要养猪国家流行的血清型为4型、5型、12型和13型,是严重危害养猪业发展的主要细菌性病原之一。目前国内还没有检测该菌抗体的商品化试剂盒。【目的】研制一种针对副猪嗜血杆菌的快速、敏感和特异性强的抗体检测试剂盒,为格拉瑟氏病的有效防控提供技术支持。【方法】表达并纯化OppA、DppA、HbpA 3种不同的副猪嗜血杆菌ABC转运体周质底物结合蛋白,使用副猪嗜血杆菌阴、阳性参考血清筛选以上3种蛋白中免疫反应性和反应特异性好的蛋白。以筛选的蛋白为包被抗原,建立检测HPS抗体的间接ELISA方法,优化间接ELISA反应条件,并组装试剂盒。在此基础上,确定该试剂盒的敏感性、特异性;通过对不同时间、不同猪场采集的2 000份临床猪血清样本进行检测,评价该试剂盒的实用性;在以上临床血清样本中随机选取200份,分别以研制的试剂盒、间接血凝试验以及进口商品化试剂盒进行检测,比较检测结...     

植物叶片葡萄糖活体检测双通道传感器的制备及应用

【目的】制备植物叶片葡萄糖活体检测传感器,以实时检测植物体叶片的葡萄糖浓度,为了解霜霉病侵染黄瓜的抗病状态提供支持。【方法】采用双通道丝网印刷电极为基底电极,使用1 mg/mL氯金酸溶液在传感器表面生成金纳米颗粒用于提高其导电性及活性面积,将5 mg/mL壳聚糖和180 mg/mL葡萄糖氧化酶混合以提高酶的固载量和活性,滴加10%的Nafion水溶液以增加对阳离子的选择性并提高电极稳定性。在此基础上研制出一种适用于活体检测植物扁平叶片中葡萄糖浓度的双通道电化学传感器。采用扫描电镜、X射线能谱、红外光谱、循环伏安法及电化学阻抗谱,对所制备传感器进行形貌、结构及电化学的表征;基于优化条件,对所制备的葡萄糖传感器进行性能测试;进一步使用黄瓜果汁进行传感器的加标试验,并应用该传感器对正常与带菌斑黄瓜叶片的葡萄糖浓度进行测定和比较。【结果】所制备植物叶片葡萄糖活体检测传感器的形貌、结构及电化学表征结果显示电极制备成功,其对葡萄糖具有良好的催化效果,反应为扩散控制过程;所设计的植物活体叶片葡萄糖检测双通道传感器的线性检测范围为1～150 mmol/L,检出限为0.784 mmol/L。采用3种不同浓度黄瓜果汁进行加标回收试验,回收率为97.17%~103.05%,相对标准误差≤ 4.52%,表明该传感器准确可靠。应用所制备的传感器检测结果表明,在双通道检测条件下,正常黄瓜叶片和霜霉菌侵染黄瓜叶片在通道1的葡萄糖浓度分别为（8.03±0.96）和（28.71±2.03） mmol/L,在通道2的葡萄糖浓度分别为（12.15±1.46）和（25.57±1.81） mmol/L,霜霉菌侵染黄瓜叶片的葡萄糖浓度较相应正常叶片增长了257.53%（通道1之间）和110.45%（通道2之间）,表明不同生理条件下同种植物叶片的葡萄糖浓度因为霜霉菌侵染而存在显著差异（P＜0.05）;病斑区域葡萄糖浓度的检测结果还显示,通道1的葡萄糖浓度高于通道2,表明通道1检测的侵染区域抗病响应迅速,侵染区相应代谢物质会发生明显变化导致邻近区域的葡萄糖逐步出现积累。【结论】所制备的传感器能够检测植物在不同生理环境下的葡萄糖浓度,可为活体植物叶片葡萄糖的实时检测提供技术支持。     

ACQUISITION HARDWARE FOR DIGITAL IMAGING

Use of digital radiography is growing rapidly in veterinary medicine. Two basic digital imaging systems are available, computed radiography (CR) and direct digital radiography (DDR). Computed radiographic detectors use a two-step process for image capture and processing. Image capture is by X-ray sensitive phosphors in the image plate. The image plate reader transforms the latent phosphor image to light photons that are converted to an analog electrical signal. An analog to digital converter is used to digitize the electrical signal before computer analysis. Direct digital detectors provide digital data by direct readout after image capture—a reader unnecessary. Types of DDR detectors are flat panel detectors and charge coupled device (CCD) detectors. Flat panel detectors are composed of layers of semiconductors for image capture with transistor and microscopic circuitry embedded in a pixel array. Direct converting flat panel detectors convert incident X-rays directly into electrical charges. Indirect detectors convert X-rays to visible light, then to electrical charges. All flat panel detectors send a digitized electrical signal to a computer using a direct link. Charge coupled device detectors have a small chip similar to those used in digital cameras. A scintillator first converts X-rays to a light signal that is minified by an optical system before reaching the chip. The chip sends a digital signal directly to a computer. Both CR and DDR provide quality diagnostic images. CR is a mature technology while DDR is an emerging technology.     

Validation and standardization of virus neutralizing test using indirect immunoperoxidase technique for the quantification of antibodies to rabies virus

A virus neutralizing test using an indirect immunoperoxidase technique (VNT-IIP) for rabies has been developed for the titration of dog and cat serum samples in Japan. The VNT-IIP has the advantage that results obtained can be viewed by the naked eye. The purpose of this study was to validate the VNT-IIP and compare it with one of the international standard methods, the fluorescent antibody virus neutralization test (FAVNT). The VNT-IIP showed satisfactory repeatability, high analytical specificity and good accuracy. Regarding the comparison between the VNT-IIP and the FAVNT, the VNT-IIP showed good agreement (91.9%), high sensitivity (92.8%) as well as specificity (87.0%) and good correlation (r = 0.92). As described above, the validation of the VNT-IIP was satisfactory and the performances of the test proved to be equivalent to those of an international standard method.     

应用血液生化指标进行山羊产绒量的间接选择

应用６项血液生化指标进行绒山羊产绒量间接选择的研究,结果表明：除血钙外,其余５项血液生化指标作为标记性状用于绒山羊产绒量的间接选择效果优于直接选择,其中以非蛋白氮、血清无机磷的效果最佳,分别是直接选择的１．６６８３倍和１．５７６３倍。     

小鼠脑提取液中铝的毛细管电泳分析

用毛细管区带电泳法(CZE)测定了小鼠脑中的铝,在10mmol/L苄胺和40％甲醇溶液中,Al3+在7min内与其他组分分离,用间接紫外吸收法在 214nm处检测,并以外标法定量,对样品进行5次测定,RDS为2.9％,回收率为96.8％,方法简便、快速、灵敏度高,重现性好,成本低。