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目的建立酶联免疫吸附多残留同步法测定饲料中硝基呋喃类违禁药物残留的分析方法。方法采用5-硝基糠醛和对羧基苯肼反应合成含有硝基呋喃类共有结构的半抗原4-硝基呋喃甲醛-(4-羧基-苯基)-腙(4-nitrofuran formaldehyde-(4-carboxyl phenyl)hydrazon,NFHBA)。通过偶联载体蛋白牛血清白蛋白(albumin from bovine serum,BSA)后的免疫原NFHBA-BSA免疫新西兰大白兔,成功制备了特异性识别呋喃环位点硝基的多克隆抗体。结果该抗体对4种目前主要使用的硝基呋喃类抗生素药物:呋喃唑酮、呋喃它酮、呋喃西林、块喃妥因的检测半抑制浓度(half maximal inhibitory concentration,IC50)分别达5.2、64、26.6、4.2 ng/mL,饲料样品平均添加回收率均在80%-90%之间。结论该方法能达到产品检测要求,可用于饲料中硝基呋喃类药物多残留同步快速检测。 相似文献
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以杀菌剂百菌清为起始反应物,通过两步化学反应合成百菌清的衍生物(半抗原);采用活化酯法与混合酸酐法分别将半抗原与载体蛋白(牛血清蛋白、卵清蛋白)进行偶联,制备百菌清的人工抗原,同时通过免疫新西兰大白兔获得抗血清,用酶联免疫吸附法(ELISA)测定其效价。利用红外光谱(IR)、质谱(MS)、核磁共振波谱(NMR)对百菌清半抗原进行表征,并用紫外扫描的方法对人工抗原进行鉴定。结果表明,成功制备出百菌清半抗原及人工抗原,并且半抗原与载体蛋白的偶联比分别为27.86:1 和12.32:1,抗血清效价约为1:1.28 × 104。 相似文献
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以苯并芘(benzo(a)pyrene,BaP)为起始反应物,通过碘代、Heck偶联两步化学反应合成苯并芘的羰基衍生物,再经肟化反应将羰基羧基化,采用活化酯法将羧基化半抗原与载体蛋白(牛血清蛋白)进行偶联,制备苯并芘的人工抗原。采用红外光谱(infrared spectroscopy,IR)、核磁共振波谱(nuclear magnetic resonancespectrum,NMR)和质谱(mass spectrometer,MS)对苯并芘中间产物进行表征,并用紫外扫描的方法对人工抗原进行鉴定。本实验成功羧化了苯并芘,并成功构建人工抗原,苯并芘与载体蛋白的偶联比为28∶1。 相似文献
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SPA用于丝素膜的生物改性研究 总被引:3,自引:0,他引:3
用生长因子RGD半抗原(GLY-ARG-GLY-ASP-SER-PRO-LYS)连接到卵清蛋白Ovalbumin(OVA)载体上诱发出了抗RGD抗体IgG,并用丝素溶液包埋SPA(Staphylo-coccal protein A,简称A蛋白或SPA)制成不溶性SPA丝素膜为材料,然后用诱发出的RGD抗体IgG结合到不溶性SPA丝素膜的表面,制成IgG-SPA丝素膜,再在其上结合粘附生长因子RGD,制成RGD-IgG-SPA丝素膜。利用这一丝素膜培养血管内皮细胞(Vas-cular Endothelial Cell,简称EC细胞),用四甲基偶氮些盐比色方法(MTT法)检测细胞的生长增殖情况。结果表明,RGD-IgG-SPA丝素膜能有效促进EC细胞的生长。对不溶性SPA丝素膜和IgG以及IgG和RGD之间的生物结合力测定,表明其结合力远大于离解力。同时在细胞培养液中没有检测到丝素膜中SPA的渗漏。RGD-IgG-SPA丝素膜为其作具有的这些优良性质为血管支架打下了基础。 相似文献
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Lan Chen Yan Zhang Karin Pacheco Shaodong Dai 《International journal of molecular sciences》2021,22(5)
Nickel (Ni2+) is one of the most common allergens, affecting around 10–15% of the general population. As the demand for orthopedic implant surgery rises, the number of surgical revisions due to joint implant failure also increases. There is evidence that some patients develop joint failure due to an immune response to a component of the implant, and we have found that Ni2+ is an especially important cause. Hence, understanding the mechanisms by which Ni2+ allergy induces joint implant failure becomes a critical research question. The structural basis of Ni2+ activation of pathogenic T cells is still not clear. The purpose of this study was to characterize Ni2+-reactive T cell repertoires derived from the peripheral blood of joint failure patients due to Ni2+ sensitization using single-cell sequencing techniques. We stimulated the proliferation of Ni2+ -reactive T cells from two implant failure patients in vitro, and sorted them for single-cell VDJ sequencing (10× genomics). We identified 2650 productive V-J spanning pairs. Both TCR α chains and β chains were enriched. TRBV18 usage is the highest in the P7 CD4+ population (18.1%), and TRBV5-1 usage is the highest in the P7 CD8+ population (12.1%). TRBV19 and TRBV20-1 segments are present in a high percentage of both P7 and P9 sequenced T cells. Remarkably, the alpha and beta chain combination of TRAV41-TRBV18 accounts for 13.5% of the CD4+ population of P7 patient. Compared to current Ni specific T cell repertoire studies of contact dermatitis, the Vα and Vβ usages of these joint implant failure patients were different. This could be due to the different availability of self-peptides in these two different tissues. However, TRBV19 (Vβ17) was among frequently used TCR β chains, which are common in previous reports. This implies that some pathogenic T cells could be similar in Ni2+ hypersensitivities in skin and joints. The alignment of the TCR CDR3β sequences showed a conserved glutamic acid (Glu) that could potentially interact with Ni2+. The study of these Ni2+ specific TCRs may shed light on the molecular mechanism of T cell activation by low molecular weight chemical haptens. 相似文献
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目的:通过人工抗原的构建,制备抗羟基红花黄色素A(HSYA)的特异性抗血清.方法:采用直接合成法将HSYA与牛血清白蛋白(BSA)和卵清蛋A(OVA)的共价偶联,分别合成免疫抗原HSYA-BSA和包被抗原HSYA-OVA,并进行相关鉴定:用合成的人工抗原免疫家兔,制得抗羟基红花黄色素A的特异性抗血清,采用酶联免疫法鉴定.结果:根据建立的标准曲线方程计算得HSYA和BSA的结合比是50:1,HSYA和OVA的结合比是112.8:1:理想的包被抗原的浓度为4μg/ml,抗血清和二抗的工作浓度分别为1:200和1:2000;交叉反应实验表明,该抗体与芦丁、金合欢素的反应率均接近于零;亲和性实验结果表明,在0.25~2μg/ml浓度范围内,抑制率与浓度呈线性关系,线性回归方程为y=-16.2131gx+102.47(r=0.9575),抑制中浓度IC<,50>为1.5μg/ml,最低检测限为0.250μg/ml.结论:通过直接偶联法可以成功制备出具有抗羟基红花黄色素A的特异性抗血清. 相似文献