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目的 体外克隆表达肾综合征出血热病毒部分核蛋白编码基因，筛选病毒核蛋白在人体体液应答中的主要抗原决定簇区，以期获得高质量核蛋白抗原，为研制新型ＨＦＲＳ基因工程诊断试剂奠定基础。方法 利用聚合酶链反应（ＰＣＲ）和限制性内切酶酶切的方法分离汉滩病毒７６－ １１８ Ｓ片段部分编码基因（ＳＡ、ＳＢ和ＳＣ），并分别克隆入Ｔ７ 原核表达载体，在大肠杆菌中表达；免疫印迹试验和酶联免疫吸附反应分析重组抗原活性。结果 截短核蛋白得到有效表达，分子量分别约为４．５ＫＤ（ｒＳＡ）、２５ｋＤ（ｒＳＢ）和１５．５ｋＤ（ｒＳＣ），Ｗｅｓｔｅｒｎ－ ｂｌｏｔ分析表明２５ｋＤ截短核蛋白具有较好的抗原活性；粗提抗原Ｄｏｔ－ＥＬＩＳＡ 方法检测ＨＦＲＳ患者血清结果与ＩＦＡ 法一致；２５ｋＤ重组小片段抗原与抗ＨＦＲＳＶ单克隆抗体５Ｈ５、Ｂ１１ 及Ｈ７可发生特异性反应。结论 汉滩病毒核蛋白的主要抗原决定簇主要集中在氨基端；原核表达的截短核蛋白在ＨＦＲＳＶ感染的血清学诊断中具有一定的价值
Objective To clone and express the partial nucleoprotein gene of hemorrhagic fever with renal syndrome virus in vitro and screen the major antigenic determinants in human humoral response in order to obtain high quality nuclear protein antigens and lay the foundation for the development of a novel diagnostic reagent for HFRS . Methods The coding genes (SA, SB and SC) of Hantaan virus 76-118 S fragment were isolated by polymerase chain reaction (PCR) and restriction endonuclease digestion and cloned into T7 prokaryotic expression vector respectively. Bacillus expression; Western blot assay and enzyme-linked immunosorbent assay for analysis of recombinant antigen activity. Results The truncated nucleoprotein was efficiently expressed with molecular weights of about 4.5KD (rSA), 25kD (rSB) and 15.5kD (rSC), respectively. Western blot analysis showed that the 25kD truncated nuclear protein had good antigenic activity. Anti-Dot-ELISA method to detect HFRS serum results consistent with the IFA method; 25kD recombinant small fragment antigen and anti-HFRSV monoclonal antibodies 5H5, B11 and H7 can react specifically. Conclusion The major epitopes of Hantaan virus nucleoprotein are mainly located at the amino terminus. The prokaryotic expression of truncated nucleoprotein is of value in the serological diagnosis of HFRSV infection