全自动化学发光免疫分析仪运行原理.ppt

1、美国雅培i2000化学发光免疫分析仪,吉林市中心医院 滕彦玲,化学发光基本原理,反应体系中的某种物质的分子(反应物、产物、中间体或荧光物质) 吸收了反应所释放的能量而由基态跃迁至激发态，然后再从激发态返回基态，同时将能量以光辐射的形式释放出来，产生化学发光. 将化学发光反应应用于分析化学，根据某一时刻的发光强度或反应的发光总量来确定体系的相应组分含量的分析方法叫化学发光分析法.,化学发光的反应必须具备两个条件,一、是该反应能释放出一定的能量，且释放出的能量可以被某种反应产物或中间体所吸收，使之处于激发态； 二、是这种激发态产物应具有一定的化学发光量子产率，或者可以将其能量有效地转移给某种荧光物

2、质，产生光辐射. 基于此，并不是任何化学反应都能产生化学发光反应，因此，如果测量条件适当，化学发光分析会有足够的选择性.,化学发光现象的发现,最早发现的化学发光现象发生在生物体内，即荧火虫，现在称之为生物发光(Bioluminescence). 到了十九世纪后期人们发现简单的非生物有机化合物也能产生化学发光. 1877年，发现洛汾碱(2，4，5-三苯基咪唑)在碱性介质中被过氧化氢等试剂氧化时发出绿色的光. 1928年，观察到鲁米诺(3-氨基苯二甲酰肼)在碱性介质中的 化学发光行为. 1935年，第一个报告了光泽精(N,N-二甲基二吖啶硝酸盐)与过氧化氢反应产生化学发光. 到现在的吖啶酯、三联吡

3、啶钌等发光标记物应用技术的成熟。,化学发光分析法的特点,由于化学发光分析不使用任何光源，避免了背景光和杂散光的干扰，降低了噪声，大大提高了信噪比，因而，化学发光分析法一般都有很高的灵敏度，通常可测定纳克级或皮克级的化学成分. 测量化学发光强度，多采用光电转换装置，用函数记录仪或数显装置，记录化学发光的相对强度. 以最大发光强度(曲线的峰高)定量被测组分的浓度. 由于流动注射分析(FIA)技术的发展，流动注射化学发光仪也广泛使用. 配备微机系统，自动进样，储存记录，打印结果，使化学发光分析的速度更快.,化学发光的分类,化学发光反应参与的免疫测定分为二种类型。 第一种是以发光剂作为酶免疫测定的底物

4、，通过发光反应增强测定的敏感性。 如：化学发光酶免疫测定（CLEIA，与酶标ELISA法类似，即最后一步酶反应所用底物为发光剂 ） 第二种是以发光剂作为抗体或抗原的标记物，直接通过发光反应检测标本中抗原或抗体的含量。 如：化学发光标记免疫测定亦称化学发光免疫测定（CLIA）、电化学发光免疫测定（ECLI）,ABBOTT i2000SR 结构,1. i2000SR processing module 2. RSH (retest sample handler) 3. SCC (system control center),ABBOTT i2000SR 结构,1. Front processing

5、 center cover 2. Processing module keypad 3. Supply and waste center door 4. Card cage door,1. Rear processing center cover 2. Rear processing center access panel 3. Power supply panel 4. Pump bay panel,ABBOTT i2000SR 结构,Sample hardware components: Provide sample aspiration and dispense. 2. Reagent

6、hardware components: Provide reagent aspiration and dispense. 3. Process path hardware components: Position the RVs for sample and reagent aspiration, mixing, washing, and CMIA processing,1. Sample pipettor,STAT pipettor,Sample and STAT pipettors (S and ST): 2. Sample and STAT syringes (SS and STS):

7、 3. Sample and STAT wash stations (SW and STW):,Reagent carousel 2. Reagent bar code reader 3. Reagent pipettors 4. Reagent syringes 5. Reagent wash stations,1. Load diverter 2. RV access door 3. RV loader and hopper assembly 4. STAT diverter5. Vortexers (Mix)6. Wash zone diverter 7. Wash zone manif

8、olds (WZ1, WZ2):8. Process path drive motor 9. Pre-trigger/trigger manifold 10. CMIA reader (CMIA)11. Liquid waste arm 12. RV unloader,ABBOTT i2000SR检测原理,Architect I系统采用化学发光微粒子免疫分析技术（Chemiluminesent Microparticle ImmunoAssay, CMIA）检测样品中的抗原，抗体和分析物。 即：磁性微粒子包被的捕捉分子（抗原，抗体或病毒颗粒）特异的与被分析物中抗原、抗体结合形成抗原抗体复合物，

9、再与吖啶酯标记的连接物反应形成双抗体夹心抗原抗体复合物。吖啶酯在过氧化氢的稀碱溶液中发生氧化还原反应生成10-甲基吖啶酮，当它恢复到基态时发光，根据发光强度可计算出分析物的浓度 。,ABBOTT i2000SR试剂组成,试剂组成 中圈（RI）：包被有抗体的顺磁性微粒子 内圈（R2）：吖啶酯包被的抗体 外圈：样本稀释液 预激发液：1.32%的过氧化氢 激发液：0.35mol./L的氢氧化钠 清洗缓冲液：磷酸盐缓冲液,R1试剂,R2试剂,ABBOTT i2000SR反应类型,1.双抗原（抗体）夹心一步法 2.双抗原（抗体）夹心两步法 Assay processing for One step 25

10、 Assay processing for Two step 18-4 STAT assay processing for One step 11 STAT assay processing for Two step 4-4,Assay processing for Two step 18-4 (iSystem),1. At position 1 the sample pipettor dispenses the sample into the RV (reaction vessel). 2. At position 2 the R1 pipettor dispenses the microp

11、articles. 3. At position 3 the vortexer mixes the sample and microparticles. 4. At positions 4 - 63 the reaction mixture incubates for 18 minutes.,共112个RV杯空位，每18秒转一个空位，200T/h。常规28Min/TEST,Assay processing for Two step 18-4 (iSystem),5. At positions 64 - 67 the wash zone 1 manifold washes the reactio

12、n mixture in the RV, and then removes unbound materials,Assay processing for Two step 18-4 (iSystem),6. At position 71 the R2 pipettor dispenses acridinium-labeled conjugate. 7. At position 72 the vortexer mixes the reaction mixture. 8. At positions 73 - 86 the reaction mixture incubates for 4 minut

13、es.,Assay processing for Two step 18-4 (iSystem),9. At positions 87 - 90 the wash zone 2 manifold washes the reaction mixture in the RV, and then removes unbound materials.,Assay processing for Two step 18-4 (iSystem),10. At position 94 the pre-trigger nozzle dispenses Pre-Trigger Solution to the re

14、action mixture, and then mixes using the vortexer. 11. At position 98 the CMIA optical system takes a background read, the trigger nozzle dispenses Trigger Solution to the reaction mixture, and then the CMIA optical system takes an activated read. 12. At position 100 the liquid waste arm aspirates t

15、he liquid waste from the RV. 13. At position 109 the RV unloader removes the RV,Assay processing for One step 25 (iSystem),1. At position 1 the sample pipettor dispenses the sample into the RV (reaction vessel). 2. At position 2 the R1 pipettor dispenses the microparticles and acridinium-labeled con

16、jugate. NOTE: For a delayed one-step assay the R2 pipettor adds the acridinium-labeled conjugate at position 71 and the vortexer mixes the reaction mixture at position 72. 3. At position 3 the vortexer mixes the sample, microparticles, and conjugate. 4. At positions 4 - 86 the reaction mixture incub

17、ates for 25 minutes.,Assay processing for One step 25 (iSystem),5. At positions 87 - 90 the wash zone 2 manifold washes the reaction mixture in the RV, and then removes unbound materials. Next position is same to the two step18-4.,STAT assay processing for One step 11 (iSystem),1. At position 47 the

18、 STAT pipettor dispenses the sample into the RV (reaction vessel). 2. At position 48 the R2 pipettor dispenses the microparticles and acridinium-labeled conjugate. 3. At position 49 the vortexer mixes the sample, microparticles, and conjugate. 4. At positions 50 - 86 the reaction mixture incubates f

19、or 11 minutes.,STAT assay processing for One step 11 (iSystem),5. At positions 87 - 90 the wash zone 2 manifold washes the reaction mixture in the RV, and then removes unbound materials. Next position is same to the two step18-4.,STAT assay processing for Two step 4-4 (iSystem),1. At position 47 the

20、 STAT pipettor dispenses the sample into the RV (reaction vessel). 2. At position 48 the R2 pipettor dispenses the microparticles. 3. At position 49 the vortexer mixes the sample and microparticles. 4. At positions 50 - 63 the reaction mixture incubates for 4 minutes.,STAT assay processing for Two step 4-4 (iSystem),5. At positions 64 - 67 the wash