血清抗Ⅱ型胶原蛋白抗体检测法的建立与应用  

Establishment of enzyme-linked immunosorbent assay for anti-collagen type II antibodies and its clinical significance in rheumatoid arthritis

作　　者：姜旭淦[1] 陈盛霞[1] 吴琴[1] 汤俊民 马建芬[2] 储青 吴亮[1] 颜小云[1] 刘荻 姚加平 许化溪[1] 

机构地区：[1]江苏大学基础医学与医学技术学院,江苏镇江212001 [2]宜兴市人民医院,江苏宜兴214200 [3]灌云县人民医院检验科,江苏灌云222000

出　　处：《临床检验杂志》2008年第3期176-178,共3页Chinese Journal of Clinical Laboratory Science

基　　金：江苏省高校自然科学指导性计划项目(06KJD310036);江苏大学第六批学生科研课题立项(06A090)

摘　　要：目的建立血清抗Ⅱ型胶原蛋白抗体的ELISA法,探讨其在类风湿关节炎(RA)患者中的应用价值。方法以自提可溶性鸡Ⅱ型胶原蛋白(soluble chicken collagen type Ⅱ,SCCⅡ)作为包被抗原,优化实验条件,建立抗Ⅱ型胶原蛋白抗体间接ELISA法。对124例RA病人和54例健康人血清抗Ⅱ型胶原蛋白抗体进行检测。结果血清抗Ⅱ型胶原蛋白抗体的检测条件为:SCCⅡ包被浓度为20μg/ml;以含10%羊血清的缓冲液进行封闭、稀释血清和酶标抗体;采用双波长(450nm/630nm)进行测定。RA患者和健康人血清抗Ⅱ型胶原蛋白抗体阳性率分别为19.35%(24/124)和1.85%(1/54),两组有明显的统计学差异(P<0.01)。结论以SCCⅡ为包被抗原检测抗Ⅱ型胶原蛋白抗体可作为判断RA患者病情的一个辅助诊断指标。Objective To establish an enzyme-linked immunosorbent assay （ELISA） for anti-collagen type Ⅱ antibodies and investigate the clinical significance in rheumatoid arthritis （RA） patients. Methods The soluble chicken collagen type Ⅱ （SCCⅡ） purified from sternal cartilage was used as coating antigen, and an improved indirect ELISA method for detecting anti-collagen type Ⅱ antibodies was established by optimizing experimental conditions. Anti-collagen type Ⅱ antibodies in 124 RA patients and 54 normal individuals were detected by the established method. Results The optimized ELISA conditions for detecting serum anti-collagen type Ⅱ antibodies were as follows. The coated SCCⅡ concentration was 20 g/ml. The buffer contained 10% goat serum was used as blocking reagent and diluent for serum samples and HRP-labeled antibodies. The absorbanees were detected by dual-wavelength spectrophotometry at 450 nm/630 nm. The positive rates of anti-collagen type Ⅱ antibodies in the sera of RA patients and normal individuals were 19.35% and 1.85% respectively. The difference was very notable between the two groups （P 〈 0.01 ）. Conclusion SCCⅡ can be used as coating antigen of ELISA for detecting anti-collagen type Ⅱ antibody in serum. The buffer containing 10% goat serum was used as blockiug reagent and diluents for serum and enzyme-labeled antibody since it can prevent nonspecific adsorption. The anti-collagen type Ⅱ antibody may seine as a supplemental parameter in diagnosis of RA.

关 键 词：类风湿关节炎 酶联免疫吸附实验 Ⅱ型胶原蛋白 抗Ⅱ型胶原蛋白抗体 

分 类 号：R446.6[医药卫生—诊断学]