CAJ | 학술논문

目的研制一种用于产前筛查孕母血清游离β绒毛膜促性腺激素(free hCG β)的时间分辨荧光免疫(TRFIA)试剂盒.方法采用2株free hCG β单克隆抗体(McAb),一株用于固相包被(3 μg/ml),另一株用于标记铕(Eu~(3+))制备Eu~(3+)-free hCG β McAb,以β-萘甲酰三氟丙酮为主要成分的增强液,应用双抗体夹心法建立free hCG β TRFIA法,并对其进行性能评价.采用线性回归分析及配对t检验进行统计学分析.结果该试剂盒的剂量-反应曲线线性相关系数(r)达0.9990;分析内、分析间变异系数(CV)均小于10.00%;标准品各点实测值与标示值的偏差(DIFF)均小于5.00%;灵敏度不高于0.20 μg/ml;以国家标准品为对照,标准品效价比在0.900～1.100间;与人促甲状腺激素(hTSH)、人黄体生成素(hLH)、人卵泡刺激素(hFSH)和人绒毛膜促性腺激素(hCG)交叉反应低;线性范围为2.00～200 ng/ml;与同类国外试剂比对,线性回归方程为Y=x+0.0565,r=0.9771,检测结果间差异无统计学意义(t=0.1839,P>0.05);国内外同类仪器比对检测,线性回归方程为y=0.9894x+0.0957,r=0.9995,检测结果间差异无统计学意义(t=-0.9669,P>0.05);室间质量评价实测值与靶值偏倚(-11.93%～7.01%)符合标准;干扰实验表明乙二胺四乙酸钾(EDTA-K_2)>0.54 mmol/L、柠檬酸钠≥1.06 mmol/L、草酸钾(K_2C_2O_4H_2O)≥2.12 mmol/L、氟化钠(NaF)≥29.76 mmoL/L时,自制试剂盒检测结果明显偏低(干扰程度均>10%),肝素≤150 U/L、血红蛋白≤12 g/L、三酰甘油≤21.54 mmol/L和胆红素≤818 μmol/L对该试剂盒检测结果无明显干扰(干扰程度均<10%);温度升高会使样本检测结果偏高.结论该试剂盒具有灵敏度高、特异性强、准确度好、线性范围宽等优点,与国外的同类产品性能相仿,能满足临床需要.
목적 연제일충용우산전사사잉모혈청유리β융모막촉성선격소(free hCG β)적시간분변형광면역(TRFIA)시제합.방법 채용2주free hCG β단극륭항체(McAb),일주용우고상포피(3 μg/ml),령일주용우표기유(Eu~(3+))제비Eu~(3+)-free hCG β McAb,이β-내갑선삼불병동위주요성분적증강액,응용쌍항체협심법건립free hCG β TRFIA법,병대기진행성능평개.채용선성회귀분석급배대t검험진행통계학분석.결과 해시제합적제량-반응곡선선성상관계수(r)체0.9990;분석내、분석간변이계수(CV)균소우10.00%;표준품각점실측치여표시치적편차(DIFF)균소우5.00%;령민도불고우0.20 μg/ml;이국가표준품위대조,표준품효개비재0.900～1.100간;여인촉갑상선격소(hTSH)、인황체생성소(hLH)、인란포자격소(hFSH)화인융모막촉성선격소(hCG)교차반응저;선성범위위2.00～200 ng/ml;여동류국외시제비대,선성회귀방정위Y=x+0.0565,r=0.9771,검측결과간차이무통계학의의(t=0.1839,P>0.05);국내외동류의기비대검측,선성회귀방정위y=0.9894x+0.0957,r=0.9995,검측결과간차이무통계학의의(t=-0.9669,P>0.05);실간질량평개실측치여파치편의(-11.93%～7.01%)부합표준;간우실험표명을이알사을산갑(EDTA-K_2)>0.54 mmol/L、저몽산납≥1.06 mmol/L、초산갑(K_2C_2O_4H_2O)≥2.12 mmol/L、불화납(NaF)≥29.76 mmoL/L시,자제시제합검측결과명현편저(간우정도균>10%),간소≤150 U/L、혈홍단백≤12 g/L、삼선감유≤21.54 mmol/L화담홍소≤818 μmol/L대해시제합검측결과무명현간우(간우정도균<10%);온도승고회사양본검측결과편고.결론 해시제합구유령민도고、특이성강、준학도호、선성범위관등우점,여국외적동류산품성능상방,능만족림상수요.
Objective To promote the development of the prenatal screening, and to develop a time-resolved fluoroimmunoassay (TRFIA) for maternal serum free human chorionic gonadotropin β subunit (free hCG β) as an aid in prenatal screening, and to prepare its test reagent. Methods Solid-phase sand-wich TRFIA was used for quantification of free hCG β, which involved two anti-free hCG β monoclonal anti-bodies (McAb). One anti-free hCG β McAh was coated on microplates, the other was labeled by Europi-um. The main component of enhancement solution was 2-naphthalene trifluoroacetone. The assay methodolo-gy was evaluated. Linear regression analysis and paired samples t-test were used. Results The correlation coefficient (r) of self-made kit dose-response curve reached 0.9990. The intra-assay and inter-assay coeffi-cient of variation (CV) was lower than 10.00%. The relative bias measured values of kit standard were lower than 5.00% difference from their labeled values. The sensitivity was better than 0.20 ng/ml. The ra-tio of self-made kit standard measured value over national standard value was 0.900～1.100. There was lit-tle cross-reaction with human thyroid stimulating hormone (hTSH), human luteinizing hormone (hLH),human follicle stimulating hormone (hFSH), and hCG. The linear range was 2.00 to 200 ng/ml. Com-pared with imported kit, y = x + 0.0565, r=0.9771, and the results showed no statistical significance (paired samples t-test, t =0.1839, P >0.05). The values of self-made kit tested by self-made equipment and imported equipment showed no difference (paired samples t-test, t=-0.9669, P > 0.05), y=0.9894x+0.0957, r=0.9995. The results of external quality assessment achieved 100%. EDTA-K_2 (≥ 0.54 mmol/L), sodium citrate (≥1.06 mmol/L), K_2C_2O_4H_2O (≥2.12 mmol/L) or NaF (≥29.76 mmoL/L) in the tested samples had interference in the assay (interference > 10%). Sodium heparin (≤ 150 U/L),haemoglobin (≤12 g/L), triglyceride (≤21.54 mmol/L) or bilirubin (≤818 μmol/L) in the tested samples had no significant interference in the assay (interference < 10%). The results of samples would be ascended due to the instability of hCG if the temperature was elevated. Conclusion The quality of self-made kit was in keeping with imported kit.