Production of anti-human thyroglobulin and anti-thyroid hormone antibodies in rabbits immunized with human thyroglobulin

Two rabbits (TG-1, TG-2) were immunized with human thyroglobulin (HTg) and bled serially. Antisera were obtained at different times after the first immunization and kept separately and studied. In both rabbits production of anti-HTg, and anti-thyroid hormone antibodies such as anti-thyroxine (T4) and anti-triiodothyronine (T3) antibodies was observed. Binding parameters of anti-HTg antibodies with HTg, T4, and T3 were calculated in two selected antisera (70-day and 249-day). The Scatchard's plots of these antibodies were all curve-linear and were analyzed in two components: one, higher binding constant (Ka1) and smaller binding capacity (Cap1) and the other, lower binding constant (Ka2) and larger binding capacity (Cap2). Ka1 values of anti-HTg, anti-T4, and anti-T3 antibodies in sera from TG-1 obtained from 70-day and 249-day bleeding were 1.1 X 10(10) M-1, 6.0 X 10(9) M-1. 7.9 X 10(8) M-1 and 1.7 X 10(10) M-1, 6.5 X 10(9) M-1, 1.0 X 10(9) M-1, respectively. Those from TG-2 were 1.7 X 10(10) M-1, 1.8 X 10(9) M-1, 6.4 X 10(8) M-1 and 2.0 X 10(10) M-1, 3.1 X 10(9) M-1, 1.6 X 10(9) M-1, respectively. The significance of the production of anti-HTg and anti-thyroid hormone antibodies in rabbits immunized with HTg in relation to the antigenic structure of HTg molecule was discussed.     

Production of anti-human thyroglobulin and anti-thyroid hormone antibodies in rabbits immunized with homogenate of human papillary cancer tissue

Papillary cancer tissue of the thyroid gland removed from each of three patients was homogenized in phosphate buffer followed by centrifugation. Each of three rabbits was immunized with each of the supernatants (TC-1, TC-2, TC-3). These rabbits were immunized on days 0, 7, 14, and 21, and serum from each rabbit, obtained 4 weeks after the first immunization, was examined for the presence of anti-human thyroglobulin (HTg), anti-thyroxine (T4), and anti-triiodothyronine (T3) antibodies. Production of anti-HTg antibodies was observed in all three rabbits. In addition, despite the low content of iodine, T3, and T4 in thyroglobulin that had been purified from the papillary cancer tissues (p-HTg), production of anti-T4 and anti-T3 was observed in two of the three rabbits, and the other immunized with TC-1 showed anti-T4 but no anti-T3 antibodies. The significance of the production of anti-thyroid hormone antibodies in rabbits with respect to the antigenic structure of p-HTg with low content of iodine and thyroid hormone is discussed.     

Wnt10a在甲乳癌及桥本合并甲乳癌中的差异表达

目的：探讨无翅型MMTV整合位点家族成员10a（Wnt10a）在甲状腺乳头状癌(papillary thyroid carcinoma,PTC)和桥本合并 甲状腺乳头癌（Hashimoto''s thyroiditis associated with papillary thyroid carcinoma）中的表达及其临床意义。方法：采用qRT-PCR方 法检测Wnt10a mRNA 在41 例新鲜甲状腺组织（PTC 组织18 例,HT 合并PTC 组织12 例,结节性甲状腺肿组织11 例）中的表 达;用免疫组化SP法检测Wnt10a 蛋白在50 例甲状腺石蜡标本（PTC 组织20 例,HT合并PTC组织20 例,结节性甲状腺肿组织 10 例）中的表达。结果：① RT-PCR Wnt10a mRNA 在甲状腺癌组织中的表达明显高于其在良性甲状腺肿瘤组织中的表达(P < 0.01) ,差异有显著性;Wnt10a mRNA在PTC、HT合并PTC组织中的阳性表达强度分别为2.49× 10-5（0.68× 10-5,15.28× 10-5）, 1.26× 10-（5 0.97× 10-5,13.73× 10-5）;P<0.05,差异具有显著性。Wnt10a mRNA 在PTC、HT 合并 PTC 组织中的表达分别是结节性甲 状腺肿组织的9.98、2.69 倍。②SP 法：Wnt10a 在甲状腺癌组织中的表达明显高于其在良性甲状腺肿瘤组织中的表达。Wnt10a mRNA 和蛋白在甲状腺癌组织中的表达与肿瘤大小、临床分期、淋巴结转移等临床因素无明显关系(P>0.05),只与甲状腺癌组织 的病理分型相关,（P<0.05)。结论：Wnt10a 在甲状腺乳头状癌和桥本合并甲状腺乳头癌中呈现高表达,且有差异性,有望作为甲状 腺癌的病理分型的参考指标,有助于甲状腺癌的早期诊断及病理分型的判定,使患者得到及时有效治疗。     

Production of the polyclonal anti-human metallothionein 2A antibody with recombinant protein technology

Metallothionein 2A(MT2A)is a small stress response protein that can be induced by exposureto toxic metals.It is highly expressed in breast cancer cells.In this study,the cDNA encoding the humanMT2A protein was expressed as glutathione S-transferase(GST)fusion protein in Escherichia coli.Recombinant MT2A proteins were loaded onto 12% sodium dodecylsulfate-polyacrylamide gel and separatedby electrophoresis,the recombinant protein was visualized by Coomassie blue staining and the 33 kDarecombinant GST-MT2A fusion protein band was cut out from the gel.The gel slice was minced and used togenerate polyclonal antisera.Immunization of rabbit against MT2A protein allowed the production of hightiter polyclonal antiserum.This new polyclonal antibody recognized recombinant MT2A protein in Westernblot analysis.This low-cost antibody will be useful for detection in various immuno-assays.     

噬菌体展示技术构建和表达抗人整合素β1单链Fv抗体

整合素（Integrins）是细胞膜蛋白的组成成分,它们是一类细胞粘附分子,并与形态形成的调节密切相关。整合素在细胞与细胞之间,细胞与基质之间的粘附中发挥着重要的作用。本研究通过选择使用鼠抗人整合素β1抑止性单克隆抗体和刺激性单克隆抗体,利用噬菌体展示,成功地构建了单链Fv抗体（scFv）,并对其与整合素β1的结合,选择最佳的可溶性抗体的生产条件等诸方面,进行了应用性研究,同时,也应用链转换（Chain shuffling）试图改变抗体亲和力进行了探索性的试验。