中华核医学杂志
中華覈醫學雜誌
중화핵의학잡지
CHINESE JOURNAL OF NUCLEAR MEDICINE
2009年
1期
50-54
,共5页
黄干%李璋巍%金河来%王霞%王建平%周智广
黃榦%李璋巍%金河來%王霞%王建平%週智廣
황간%리장외%금하래%왕하%왕건평%주지엄
放射免疫测定%胰岛素抗体%糖尿病,胰岛素依赖型%糖尿病,非胰岛素依赖型
放射免疫測定%胰島素抗體%糖尿病,胰島素依賴型%糖尿病,非胰島素依賴型
방사면역측정%이도소항체%당뇨병,이도소의뢰형%당뇨병,비이도소의뢰형
Radioimminoassay%Insulin antibodies%Diabetes mellitus,insulin-dependent%Diabtes mellitus,non-insulin-dependent
目的 建立胰岛素自身抗体(IAA)的微量平板放射免疫(RIA)法,并探讨其临床应用价值.方法 125Ⅰ-胰岛素与血清在微量平板4℃保温72 h后,免疫复合物转移至已包被蛋白A的Millipore过滤平板中并洗涤,于多功能液体闪烁发光分析仪上测计数.以IAA指数≥0.06作为阳性标准,通过参加第四次国际糖尿病自身抗体检测标准化评估(DASP2005),评价方法的灵敏度与特异性.并检测71例1型糖尿病(T1DM)患者、551例初诊2型糖尿病(T2DM)患者以及317名健康对照者IAA水平.采用SPSS 11.5软件进行t检验、非参数检验x2检验和直线相关分析;一致率用Kappa值评估.结果 (1)优化的检测条件为5 ul血清与2×104计数·min-1的125Ⅰ-胰岛素于4℃缓慢振摇保温72 h.(2)该方法批内CV4.8%～8.9%,批间CV6.4%～10.5%,DASP 2005反馈结果显示该法诊断T1DM灵敏度50%(25/50),特异性97%(97/100).与国产IAA RIA试剂盒进行比较,结果判定总体一致率为72.9%(Kappa值0.402),相关系数(r)为0.678(P<0.001);26例结果不相符的标本中25例为该法检测阳性而国产RIA试剂盒检测阴性.(3)该方法检测TIDM患者的阳性率为19.7%(16/71),显著高于健康对照组的0.9%(3/317,x2=54.36,P<0.001),其中0～9岁T1DM组9例中IAA阳性5例.检测初诊T2DM患者IAA阳性率1.5%,与健康对照组差异无统计学意义(x2=0.95,P>0.05).结论 IAA微量平板RIA法灵敏度与特异性好,可应用于临床,IAA在婴幼儿T1DM患者中阳性率较高.
目的 建立胰島素自身抗體(IAA)的微量平闆放射免疫(RIA)法,併探討其臨床應用價值.方法 125Ⅰ-胰島素與血清在微量平闆4℃保溫72 h後,免疫複閤物轉移至已包被蛋白A的Millipore過濾平闆中併洗滌,于多功能液體閃爍髮光分析儀上測計數.以IAA指數≥0.06作為暘性標準,通過參加第四次國際糖尿病自身抗體檢測標準化評估(DASP2005),評價方法的靈敏度與特異性.併檢測71例1型糖尿病(T1DM)患者、551例初診2型糖尿病(T2DM)患者以及317名健康對照者IAA水平.採用SPSS 11.5軟件進行t檢驗、非參數檢驗x2檢驗和直線相關分析;一緻率用Kappa值評估.結果 (1)優化的檢測條件為5 ul血清與2×104計數·min-1的125Ⅰ-胰島素于4℃緩慢振搖保溫72 h.(2)該方法批內CV4.8%～8.9%,批間CV6.4%～10.5%,DASP 2005反饋結果顯示該法診斷T1DM靈敏度50%(25/50),特異性97%(97/100).與國產IAA RIA試劑盒進行比較,結果判定總體一緻率為72.9%(Kappa值0.402),相關繫數(r)為0.678(P<0.001);26例結果不相符的標本中25例為該法檢測暘性而國產RIA試劑盒檢測陰性.(3)該方法檢測TIDM患者的暘性率為19.7%(16/71),顯著高于健康對照組的0.9%(3/317,x2=54.36,P<0.001),其中0～9歲T1DM組9例中IAA暘性5例.檢測初診T2DM患者IAA暘性率1.5%,與健康對照組差異無統計學意義(x2=0.95,P>0.05).結論 IAA微量平闆RIA法靈敏度與特異性好,可應用于臨床,IAA在嬰幼兒T1DM患者中暘性率較高.
목적 건립이도소자신항체(IAA)적미량평판방사면역(RIA)법,병탐토기림상응용개치.방법 125Ⅰ-이도소여혈청재미량평판4℃보온72 h후,면역복합물전이지이포피단백A적Millipore과려평판중병세조,우다공능액체섬삭발광분석의상측계수.이IAA지수≥0.06작위양성표준,통과삼가제사차국제당뇨병자신항체검측표준화평고(DASP2005),평개방법적령민도여특이성.병검측71례1형당뇨병(T1DM)환자、551례초진2형당뇨병(T2DM)환자이급317명건강대조자IAA수평.채용SPSS 11.5연건진행t검험、비삼수검험x2검험화직선상관분석;일치솔용Kappa치평고.결과 (1)우화적검측조건위5 ul혈청여2×104계수·min-1적125Ⅰ-이도소우4℃완만진요보온72 h.(2)해방법비내CV4.8%～8.9%,비간CV6.4%～10.5%,DASP 2005반궤결과현시해법진단T1DM령민도50%(25/50),특이성97%(97/100).여국산IAA RIA시제합진행비교,결과판정총체일치솔위72.9%(Kappa치0.402),상관계수(r)위0.678(P<0.001);26례결과불상부적표본중25례위해법검측양성이국산RIA시제합검측음성.(3)해방법검측TIDM환자적양성솔위19.7%(16/71),현저고우건강대조조적0.9%(3/317,x2=54.36,P<0.001),기중0～9세T1DM조9례중IAA양성5례.검측초진T2DM환자IAA양성솔1.5%,여건강대조조차이무통계학의의(x2=0.95,P>0.05).결론 IAA미량평판RIA법령민도여특이성호,가응용우림상,IAA재영유인T1DM환자중양성솔교고.
Objective Insulin autoantibody (IAA) is known to exist in sera of type 1 diabetes mellitus (T1DM) patients and pre-T1DM individuals. The aim of this study was to establish a novel microtiter plate radioimmunoassay (RIA) for IAA and evaluate its clinical value. Methods Diluted 125Ⅰ-insulin was mixed with 5 ul serum samples in a 96-well microtiter plate and then incubated for 72 h on an orbital plate shaker (4℃). The immunocomplexes were transferred to another protein a coated Millipore plate, and then the plate was washed with Tri-Buffered Saline Tween-20 (TBT) buffer. Counts per minute (CPM) was measured with liquid scintillation and luminescence counter. The positive cut-off point of IAA index was defined as ≥0.06 based on the 99-percentile of the distribution in 317 healthy individuals. The specificity and sensitivity of the assay were calculated from the samples provided by the fourth Diabetes Autoantibodies Standardization Program (DASP 2005). The IAA levels were determined in 71 T1 DM and 551 newly diagnosed type 2 diabetes (T2DM) patients, and 317 healthy controls. The t test, non-parametric test, x2 test and linear correlation analysis were performed on the data using SPSS 11.5 software. The concordance rate was estimated with Kappa value. Results (1) The optimized testing condition was described as 2×104 CPM of 125Ⅰ-insulin, 5 ul serum sample and slowly horizontal shaking for 72 h. (2) The intra-assay CV was 4.8%-8.9% and inter-assay CV was 6.4%-10.5%. Based on DASP 2005 samples, the specificity and sensitivity of the assay were 97% (97/100) and 50% (25/50), respectively. Ninety-six serum samples with different IAA levels were selected and tested to compare between our new method and a domestic IAA RIA kit. The results showed that the IAA indices from the two methods were positively correlated (r= 0.678, P<0.001). The concordance rate was 72.9 %(Kappa value=0.402). There were 25 samples with discordant results, which were positive for IAA titer using the corresponding microtiter plate RIA but negative using the novel RIA kit. (3) In TIDM group the positive rate of IAA was 19.7% (16/71), higher than the healthy controls (0.9%, x2=54.36, P<0.001). The subgroup of T1DM children (with 0-9 years) showed the highest IAA positive rate (55.6% ,x2=4.85, P<0.05). In T2DM group the frequency of IAA was 1.5% (8/551), which had no significant difference comparing with that of healthy controls (x2= 0.95, P >0.05). Conclusions Our proposed microtiter plate RIA method for IAA is highly sensitive and specific, likely to be feasible for clinical application. The frequency of IAA is high in children with T1DM.
