目的:探讨异常凝血酶原(protein induced by vitamin K absence or antagonist-Ⅱ,PIVKA-Ⅱ)与高尔基体蛋白(Golgi glycoprotein,GP73)联合检测对诊断血清甲胎蛋白(alpha-fetoprotein,AFP)低浓度的原发性肝癌(primary hepatic carcinoma...目的:探讨异常凝血酶原(protein induced by vitamin K absence or antagonist-Ⅱ,PIVKA-Ⅱ)与高尔基体蛋白(Golgi glycoprotein,GP73)联合检测对诊断血清甲胎蛋白(alpha-fetoprotein,AFP)低浓度的原发性肝癌(primary hepatic carcinoma,PHC)的临床价值。方法:选取本院2015年3月-2016年12月AFP低浓度PHC72例,其他肝病68例,同期体检正常对照组110例,分别检测PIVKA-Ⅱ(ELISA)和GP73(ELISA)、AFP(化学发光法)的血清浓度水平。结果:PIVKA-Ⅱ与GP73联合检测在AFP低浓度PHC的系列试验特异度高达100%,联合试验敏感度达90.2%。结论:单独检测PIVKA-Ⅱ和GP73对AFP低浓度肝癌具有较高的诊断价值,PIVKA-Ⅱ与GP73联合检测可提高AFP低浓度肝癌诊断。展开更多
[Objective] The aim of this study is to clone the active promoter of golgi membrane glycoprotein GP73. [Method] The sequence within the range of upstream 1 000 bp and downstream 400 bp of transcription initiation site...[Objective] The aim of this study is to clone the active promoter of golgi membrane glycoprotein GP73. [Method] The sequence within the range of upstream 1 000 bp and downstream 400 bp of transcription initiation site was analyzed, the genomic DNA of hepatoma cell line Huh-7 was regarded as template for PCR amplification. The amplified fragment was cloned into recombinant construct with enhanced green fluorescent protein (EGFP) report gene. The expression of EGFP in recombinants were observed under fluorescence microscope after transfection, with the assistant of flow cytometry. [Result] The 1 310 bp ranging between upstream 980 bp and downstream 330 bp of transcription initiation site assumed promoter function. The region contains two core promoters and several conserved sequences including TATA box and many DNA binding sites such as NF-κB, AP1, GC-SP1. [Conclusion] The cloning of the promoter provides a reference for the study of the transcription mechanism of GP73.展开更多
目的:高尔基体蛋白73(Golgi protein 73,GP73)是新近发现的肝细胞性肝癌(hepatocellular carcinoma,HCC)的重要血清学标志物。通过克隆、表达人GP73,制备抗GP73单克隆抗体并鉴定其特性。方法:克隆GP73基因,构建GP73原核表达载体pComb3XS...目的:高尔基体蛋白73(Golgi protein 73,GP73)是新近发现的肝细胞性肝癌(hepatocellular carcinoma,HCC)的重要血清学标志物。通过克隆、表达人GP73,制备抗GP73单克隆抗体并鉴定其特性。方法:克隆GP73基因,构建GP73原核表达载体pComb3XSS-GP73,诱导表达,以HisFF Trap亲和层析柱纯化GP73-6×His重组融合蛋白。以该融合蛋白免疫BALB/c小鼠,制备抗GP73单克隆抗体。以间接法ELISA检测抗体效价;Western blot鉴定抗体特异性;免疫共沉淀法初步检测肝癌患者血清中GP73表达水平。结果:成功表达并纯化了GP73-6×His重组蛋白;获得5株稳定分泌抗人GP73单克隆抗体杂交瘤细胞株;免疫共沉淀证实抗GP73单克隆抗体能与肝癌患者血清中GP73蛋白特异性结合,GP73水平在肝癌患者血清中较正常人显著升高。结论:成功制备了抗人GP73单克隆抗体,为后期建立检测人GP73的双抗体夹心ELISA方法打下基础。展开更多
文摘目的:探讨异常凝血酶原(protein induced by vitamin K absence or antagonist-Ⅱ,PIVKA-Ⅱ)与高尔基体蛋白(Golgi glycoprotein,GP73)联合检测对诊断血清甲胎蛋白(alpha-fetoprotein,AFP)低浓度的原发性肝癌(primary hepatic carcinoma,PHC)的临床价值。方法:选取本院2015年3月-2016年12月AFP低浓度PHC72例,其他肝病68例,同期体检正常对照组110例,分别检测PIVKA-Ⅱ(ELISA)和GP73(ELISA)、AFP(化学发光法)的血清浓度水平。结果:PIVKA-Ⅱ与GP73联合检测在AFP低浓度PHC的系列试验特异度高达100%,联合试验敏感度达90.2%。结论:单独检测PIVKA-Ⅱ和GP73对AFP低浓度肝癌具有较高的诊断价值,PIVKA-Ⅱ与GP73联合检测可提高AFP低浓度肝癌诊断。
文摘[Objective] The aim of this study is to clone the active promoter of golgi membrane glycoprotein GP73. [Method] The sequence within the range of upstream 1 000 bp and downstream 400 bp of transcription initiation site was analyzed, the genomic DNA of hepatoma cell line Huh-7 was regarded as template for PCR amplification. The amplified fragment was cloned into recombinant construct with enhanced green fluorescent protein (EGFP) report gene. The expression of EGFP in recombinants were observed under fluorescence microscope after transfection, with the assistant of flow cytometry. [Result] The 1 310 bp ranging between upstream 980 bp and downstream 330 bp of transcription initiation site assumed promoter function. The region contains two core promoters and several conserved sequences including TATA box and many DNA binding sites such as NF-κB, AP1, GC-SP1. [Conclusion] The cloning of the promoter provides a reference for the study of the transcription mechanism of GP73.
