CAJ | 학술논문

目的：获得大量粉尘螨重组变应原 Der f8蛋白，检测其免疫原性。方法合成粉尘螨第8组变应原（Der f8）基因，将其连接至 pET-32 a 载体，用异丙基-β-D-硫代半乳糖苷（IPTG）诱导表达重组 Der f8蛋白，圆二色谱初步检测该蛋白二级结构，以粉尘螨过敏患者血清作为一抗，经免疫印迹法（Western-blotting）方法分析 Der f8的免疫学特性。结果重组工程菌经 IPTG 诱导后，高效表达出 Der f8蛋白，为可溶性蛋白。SDS-PAGE 结果显示，表达产物分子质量约为45 ku。Der f8重组蛋白二级结构中，α螺旋34．5％，β折叠12．6％，无规则卷曲53．9％。表达产物经亲和层析纯化后，Western Blot 印迹有明显条带显示。结论纯化后的 Der f8具有较高的纯度和较强的免疫学活性，为粉尘螨过敏反应性疾病的特异性诊断和治疗及进一步的实验研究奠定了基础。
목적：획득대량분진만중조변응원 Der f8단백，검측기면역원성。방법합성분진만제8조변응원（Der f8）기인，장기련접지 pET-32 a 재체，용이병기-β-D-류대반유당감（IPTG）유도표체중조 Der f8단백，원이색보초보검측해단백이급결구，이분진만과민환자혈청작위일항，경면역인적법（Western-blotting）방법분석 Der f8적면역학특성。결과중조공정균경 IPTG 유도후，고효표체출 Der f8단백，위가용성단백。SDS-PAGE 결과현시，표체산물분자질량약위45 ku。Der f8중조단백이급결구중，α라선34．5％，β절첩12．6％，무규칙권곡53．9％。표체산물경친화층석순화후，Western Blot 인적유명현조대현시。결론순화후적 Der f8구유교고적순도화교강적면역학활성，위분진만과민반응성질병적특이성진단화치료급진일보적실험연구전정료기출。
Objective To obtain a lot of recombinant allergen Der f8 protein from Dermatopha-goides farinae,and to detect its immunogenicity.Methods The gene encoding group 8 allergen of Dermatophagoides farinae (Der f8)was synthesized and introduced into pET-32 vector.The ex-pression of recombinant Der f8 protein was induced by IPTG.The secondary structure of Der f8 was detected by circular dichroism spectrum.The immunological characteristics were detected by Western blot assay using the patient’s sera as primary antibody.Results After IPTG induction, the engineering bacteria expressed soluble recombinant Der f8 protein.SDS-PAGE showed a band at 45 ku.The secondary structure of Der f8 included 34.5% of α-helix,12.6% of β-sheet and 53.9% of random coil.After purification by affinity chromatography,Western blot detection of the expression product showed obvious strip.Conclusion The purified Der f8 has high purity and immunological activity.It can lay a foundation for the specific diagnosis,treatment and further ex-perimental studies of dust mite allergy disease.