Application of ELISA-based kit for detecting AOZ and determining its clearance in eel tissues

Furazolidone, an antibacterial drug that was once widely used in the livestock industry and aquaculture, is now prohibited in numerous countries. It is difficult to detect residual furazolidone because it is readily metabolized in animal tissues but, by using and liquid chromatography coupled with tandem mass spectrometry, its metabolite, 3-amino-2-oxazolidinone (AOZ) can be detected. Here we describe the validity of an enzyme-linked immunosorbent assay (ELISA) kit to detect AOZ in Japanese eel Anguilla japonica tissue. ELISA is capable of detecting AOZ at 1.0 μg/kg in an eel sample with excellent accuracy and precision. Our results show that ELISA is suitable for regulatory purposes and for studying the fate of AOZ residues in eel treated with furazolidone. To measure the persistence of AOZ in eel tissues, eels (1.4–6.5g) were immersed in tanks containing 2 and 10 mg furazolidone/L for 3 h, and then maintained in a tank supplying well water for the next 160 days. The half-lives of AOZ, calculated from the linear terminal part of the excretion curve, were 25.0 days in muscle and 21.6 days in liver from fish exposed to 2 mg/L furazolidone. In the eels treated with 10 mg/L furazolidone, by contrast, high levels of AOZ were detected in liver and muscle, but the half-lives of AOZ were similar to those in fish treated with 2 mg/L furazolidone. The half-lives of AOZ in eel tissues were prolonged by the condition of low water temperature.     

细胞因子Daintain/AIF-1的纯化及其单克隆抗体的制备与鉴定

由猪小肠中纯化鉴定Daintain/AIF-1,并以其为免疫原免疫BALB/c小鼠,用杂交瘤技术制备得到4株稳定分泌抗体的单克隆细胞株。对单抗效价、亚类、亲和常数进行分析,4株单抗都属于IgG1(κ)。又经West-ern blot证实,所制备的抗体能特异性与Daintain/AIF-1结合。     

Generation of mouse monoclonal antibodies specific to tilapia immunoglobulin using fish immunoglobulin/BSA complex for monitoring of the immune response in Nile tilapia Oreochromis niloticus

The simple immunoprecipitation method was used to isolate tilapia immunoglobulin (Ig) for immunization in order to produce monoclonal antibodies (MAbs) specific to tilapia Ig. First, the tilapia antiserum against bovine serum albumin (BSA) was prepared by peritoneal injection of BSA into tilapia, and the tilapia anti‐BSA antiserum was used to precipitate BSA to form the Ig/BSA immune complex. The Ig/BSA immune complex was then injected into Swiss mice for hybridoma production. After fusion, three hybridoma clones producing MAbs specific to the tilapia antibody were selected by dot blot and Western blot. All MAbs (101A, 59G, and 11A) were bound specifically to the heavy chain of immunoglobulin M (IgM). The MAbs 101A and 59G demonstrated twofold higher affinity than MAb 11A and the commercialized antibody. However, MAbs 11A could also bind to the heavy chain of IgM in Asian seabass, Lates calcarifer, as well. These MAbs can be used to monitor the immune responses of individual fish by indirect ELISA upon exposure to various antigens.     

血清法监测人工感染弓形虫病猪特异抗体的比较

应用ELISA、LAT、IHA血清试验对8头人工感染弓形虫猪的血清抗体滴度变化规律进行了监测,并对不同检测方法的结果进行了比较.结果发现,试验猪在皮下感染弓形虫RH株速殖子后,血清中的特异性抗体首次检出的时间方面,LAT(第4 d)早于ELISA(第14 d)和IHA(第20 d);检出持续时间方面,LAT(54 d)长于ELISA(44 d)和IHA(19 d);平均检出率方面,LAT(7/8)高于ELISA(6.7/8)和IHA(5/8).试验结果表明,ELISA和LAT适合于猪弓形虫病的诊断和血清学调查,LAT更适合于猪弓形虫病的早期检测.     

烟草病毒检测技术研究进展

近年来烟草病毒病发生病普遍,且日趋严重,对烟草的产量及质量影响很大,使烟草行业蒙受巨大损失。鉴于此,建立有效的病毒检测技术,对烟草病毒进行防治十分重要。综述了国内外对烟草病毒的检测技术,主要包括:指示植物法、电镜检测方法、血清学方法、分子生物学方法等。     

抗卡那霉素单克隆抗体细胞株的筛选及间接ELISA法的建立

采用碳二亚胺(EDC)法合成卡那霉素完全抗原,免疫BALB/c小鼠,通过杂交瘤技术获得3株能稳定分泌抗卡那霉素单克隆抗体的杂交瘤(5D7、6H8、7G4)。选择其中的5D7株抗体,经ELISA鉴定,细胞上清的效价达103以上,腹水效价达到5×104以上;建立了间接竞争ELISA法,通过优化,其检测限为0.5ng/mL,抗体IC50为3.7ng/mL,与妥布霉素交叉反应率为63.8%,与其他抗生素无交叉。可见ELISA具有快速、敏感、特异、简便等特点,适合于KAN残留的快速检测,具有较高的推广应用价值。     

对生和互生玉米内源激素含量的差异分析

利用ELISA方法测定近等基因系玉米(Zeamays)对生、互生玉米的不同组织和不同生育期内源细胞分裂素ZRs(zeatin riboside group)、DHZRs(dehydro zeatin riboside group)、iPAs(isopentenyladenosine group)以及内源生长素IAA(indole acetic acid)含量差异。结果表明,不同叶序玉米在不同生育期幼胚、成熟花粉和成熟叶片的内源ZRs、DHZRs、iPAs和IAA均无显著差异;不同生育期对生叶序玉米幼苗顶端分生组织和成熟苗顶端雄幼穗ZRs、DHZRs和iPAs含量显著高于同期近等基因系的互生玉米,IAA含量在两者之间无显著差异;对生叶序玉米幼苗顶端分生组织、成熟苗顶端雄幼穗CTK/IAA(CTK表示ZRs、DHZRs和iPAs含量相加)比值显著高于同期近等基因系互生叶序玉米,但在不同生育期幼胚、叶片及花粉内均无显著差异。对生叶序的形成与生长点组织细胞分裂素含量增加密切相关。     

间接竞争性ELISA检测甲胺磷残留

采用制备的兔抗血清建立了甲胺磷（methamidophos,MTP)残留的间接竞争性ELISA方法，证明了抗血清的特异性较好，并确立了测定参数：抗血清与包被抗原的最佳工作浓度分别为1：3000和1：5000；最适检测范围为0．01－0．1μg/ml；批内变异系数7．56％，批间变异系数5．70％。     

抗犬细小病毒的单克隆抗体的制备

CPV YZ株的细胞培养液经蔗糖密度梯度离心纯化后作为免疫原免疫BALB/C小鼠 ,应用杂交瘤技术筛选出 8株能稳定分泌单克隆抗体的杂交瘤细胞 ,分别命名为F5G10、3F11F4、F11G6、6C5C2、6E4G3、6C2B11、F11D6、3D9C4株 ,各株单抗均为IgG型 ;中和试验表明 :F11G6、3D9C4、6E4G3、6C5B114株单抗有中和作用 ,Dot ELISA试验证明了 8株单抗仅针对病毒抗原决定簇的空间结构。     

蔬菜中克百威残留的ELISA法测定技术研究

以合成人工抗原复合物免疫新西兰大白兔,制备了特异性的兔抗BFNH多克隆抗血清。用间接ELISA法测得抗体血清的最高效价可达1:32000。并以BFNH-卵清蛋白(OVA)复合物,作为抗原包被96孔板,建立了间接竞争ELISA法,用于蔬菜中的克百威残留分析。通过实验确定该方法的工作条件和标准曲线,并测定了抗体血清与其它各种氨基甲酸酯类农药的交叉反应率。实验结果表明,抗体血清对其它氨基甲酸酯类农药的交叉反应率均小于10%。该方法对克百威纯品的检测限为0.1μg·ml-1,线性检测范围为102～10-3μg·ml-1。该方法测定结果与高效液相色谱测定结果相一致。