本发明专利技术涉及血清腺苷脱氨酶检测技术领域，特别涉及一种血清腺苷脱氨酶检测试剂，R1中含有缓冲液，4‑氨基氨替吡啉，PNP，XOD，POD，胆红素氧化酶，抗坏血酸氧化酶，丙三醇，聚乙二醇6000，乙二醇，甘露醇，海藻糖，BSA，烷基糖苷，防腐剂；R2中含有缓冲液，腺苷，3‑羟基‑2,4,6‑三碘苯甲酸，丙三醇，聚乙二醇6000，乙二醇，甘露醇，海藻糖，BSA，烷基糖苷，防腐剂；采用新的Trinder反应色原物质3‑羟基‑2,4,6‑三碘苯甲酸，添加多种稳定剂，显著改善了试剂稳定性；通过添加胆红素氧化酶、抗坏血酸氧化酶和新型非离子表面活性剂烷基糖苷（APG），有效避免了胆红素和抗坏血酸的干扰，防止反应体系浑浊，底物稳定性、试剂的抗干扰能力均增强。
【技术实现步骤摘要】
本专利技术涉及血清腺苷脱氨酶检测
，特别涉及一种血清腺苷脱氨酶检测试剂，还涉及使用此检测试剂的检测方法。
技术介绍
腺苷脱氨酶(ADA)是人体嘌吟核苷酸代谢中的重要酶类，广泛分布于人体各组织中，其中以胸腺、脾和其他淋巴组织中含量最高，而肝、肺、肾和骨胳肌等含量较低；血清中的ADA主要来自肝脏，是反映肝损伤的敏感指标，可作为肝功能常规检查项目之一，依据该酶活性增高或降低反映肝细胞损伤和恢复程度；慢性溶血患者红细胞ADA活性显著升高；肿瘤患者血清及组织中ADA活性均升高；结核性脑膜炎患者CSF-ADA明显升高。目前，腺苷脱氨酶的测定方法有波氏显色法、氨气敏电极法、酶法、酶偶联法等；波氏显色法试剂简单、易于操作，但该法易受外源性铵根离子的影响；氨气敏电极法虽然抗干扰能力较强，但难以控制电极的响应信号和响应时间，不适于临床大批量样品的测定；酶法较灵敏、可实现自动化分析，但重复性和准确度较差；酶偶联法结果准确、简便实用，但易受抗坏血酸和胆红素等物质干扰。鉴于此，本专利技术在酶偶联法的基础上，优化反应体系，添加胆红素氧化酶和抗坏血酸氧化酶，可以有效避免胆红素和抗坏血酸的干扰，大大增强试剂的抗干扰能力，并且采用新的Trinder反应色原物质HTBA（3-羟基-2,4,6-三碘苯甲酸），显著增强了试剂的稳定性和准确度；此外，通过添加稳定剂如丙三醇、聚乙二醇6000、甘露醇、海藻糖、BSA、乙二醇等可有效提高试剂的稳定性；且优选的新型非离子表面活性剂烷基糖苷（APG）的加入可防止反应体系浑浊，增强底物的稳定性，提高试剂的抗干扰能力；该试剂操作简便快速，适用于自动化分析，是一种更加稳定、抗干扰能力强的血清腺苷脱氨酶（ADA）试剂。
技术实现思路
本专利技术的目的是提供一种用于检测血清腺苷脱氨酶（ADA）的试剂及使用该试剂检测血清腺苷脱氨酶活性的方法。该试剂盒采用酶偶联法，可以有效检测血清腺苷脱氨酶的活性，抗干扰能力强，稳定性好等优点。基本原理：ADA（腺苷脱氨酶）催化底物腺苷脱氨生成次黄嘌呤核苷，次黄嘌呤核苷再经 PNP（嘌呤核苷磷酸化酶）作用生成次黄嘌呤。次黄嘌呤在XOD（黄嘌呤氧化酶）作用下，生成尿酸和过氧化氢（H2O2）。过氧化氢(H2O2) 在过氧化物酶存在的情况下，可与3-羟基-2,4,6-三碘苯甲酸（HTBA）和4-氨基氨替吡啉（4-AA）反应产生有色醒类化合物，有色醒类化合物的生成速率与ADA（腺苷脱氨酶）的活性相关。通过测定550nm波长处吸光度的上升速率，即有色醒类化合物生成速率来测定腺苷脱氨酶的活性。本专利技术是通过以下步骤得到的：一种血清腺苷脱氨酶检测试剂，包括试剂R1和试剂R2，所述试剂R1和试剂R2的组成如下：试剂R1中含有缓冲液··························100mmol/L，4-氨基氨替吡啉··················5mmol/L，PNP（嘌呤核苷磷酸化酶）···········1.5KU/L，XOD（黄嘌呤氧化酶）···············3.1KU/L，POD（过氧化物酶）·················5KU/L，胆红素氧化酶····················3KU/L，抗坏血酸氧化酶··················3.5KU/L，丙三醇··························4ml/L，聚乙二醇6000····················5g/L，乙二醇··························6ml/L，甘露醇··························25g/L，海藻糖··························15g/L，BSA·····························1g/L，烷基糖苷（APG）···················1g/L，防腐剂··························0.5g/L；2）试剂R2的组分为：缓冲液··························100mmol/L，腺苷····························14mmol/L，HTBA（3-羟基-2,4,6-三碘苯甲酸）···5mmol/L，丙三醇··························4ml/L，聚乙二醇6000····················5g/L，乙二醇··························6ml/L，甘露醇··························25g/L，海藻糖··························15g/L，BSA·····························1g/L，烷基糖苷（APG）···················1g/L，防腐剂··························0.5g/L。所述的血清腺苷脱氨酶检测试剂，试剂R1中缓冲液为25℃，pH为8.0的HEPES缓冲液。所述的血清腺苷脱氨酶检测试剂，试剂R2中缓冲液为25℃，pH为4.0的Tris缓冲液。所述的血清腺苷脱氨酶检测试剂，所述防腐剂为NaN3。所述的血清腺苷脱氨酶检测试剂来检测血清腺苷脱氨酶含量的检测方法，使用全自动生化分析仪利用速率法进行测定，检测主波长为550nm。所述的检测方法，R1试剂和R2试剂的比例为2:1。本专利技术的有益效果：1）添加胆红素氧化酶和抗坏血酸氧化酶，可以有效避免胆红素和抗坏血酸的干扰，大大增强试剂的抗干扰能力；2）采用新的Trinder反应色原物质HTBA（3-羟基-2,4,6-三碘苯甲酸），显著增强了试剂的稳定性和准确度；3）新型非离子表面活性剂烷基糖苷（APG）的加入可防止反应体系浑浊，增强底物的稳定性，提高试剂的抗干扰能力；4）优化反应体系并添加丙三醇、聚乙二醇6000、甘露醇、海藻糖、BSA、乙二醇等多种稳定剂，可以显著改善试剂的稳定性；5）试剂的准确度和稳定性良好，抗干扰性强，使用方便，完全可以满足临床需要。附图说明图1为两种试剂的相关性曲线图，图2为两种试剂效期稳定性曲线图，图3为实施例1试剂检测方法，图4为实施例试剂抗干扰性能比较，图5为实施例1试剂与市场常见并得到认可的血清腺苷脱氨酶测定试剂盒对比检测结果。具体实施方式下面结合具体实施例对本专利技术进行进一步说明。实施例1血清腺苷脱氨酶的检测试剂，包括试剂R1和试剂R2：1）其R1的组成为：HEPES（4-羟乙基哌嗪乙磺酸）缓冲液(pH=8.0，25℃) ··100mmol/L，4-氨基氨替吡啉·································5mmol/L，PNP（嘌呤核苷磷酸化酶）··························1.5KU/L，XOD（黄嘌呤氧化酶）······························3.1KU/L，POD（过氧化物酶）································5KU/L，胆红素氧化酶···································3KU/本文档来自技高网...
【技术保护点】
一种血清腺苷脱氨酶检测试剂，其特征在于包括试剂R1和试剂R2，所述试剂R1和试剂R2的组成如下：1）试剂R1的组分为：缓冲液·························100mmol/L，4‑氨基氨替吡啉·················5mmol/L，PNP（嘌呤核苷磷酸化酶）··········1.5KU/L，XOD（黄嘌呤氧化酶）··············3.1KU/L，POD（过氧化物酶）················5KU/L，胆红素氧化酶···················3KU/L，抗坏血酸氧化酶·················3.5KU/L，丙三醇·························4ml/L，聚乙二醇6000···················5g/L，乙二醇·························6ml/L，甘露醇·························25g/L，海藻糖·························15g/L，BSA····························1g/L，烷基糖苷（APG）··················1g/L，防腐剂·························0.5g/L；2）试剂R2的组分为：缓冲液·························100mmol/L，腺苷···························14mmol/L，HTBA（3‑羟基‑2,4,6‑三碘苯甲酸）··5mmol/L，丙三醇·························4ml/L，聚乙二醇6000···················5g/L，乙二醇·························6ml/L，甘露醇·························25g/L，海藻糖·························15g/L，BSA····························1g/L，烷基糖苷（APG）··················1g/L，防腐剂·························0.5g/L。...
【技术特征摘要】
1.一种血清腺苷脱氨酶检测试剂，其特征在于包括试剂R1和试剂R2，所述试剂R1和试剂R2的组成如下：1）试剂R1的组分为：缓冲液·························100mmol/L，4-氨基氨替吡啉·················5mmol/L，PNP（嘌呤核苷磷酸化酶）··········1.5KU/L，XOD（黄嘌呤氧化酶）··············3.1KU/L，POD（过氧化物酶）················5KU/L，胆红素氧化酶···················3KU/L，抗坏血酸氧化酶·················3.5KU/L，丙三醇·························4ml/L，聚乙二醇6000···················5g/L，乙二醇·························6ml/L，甘露醇·························25g/L，海藻糖·························15g/L，BSA····························1g/L，烷基糖苷（APG）··················1g/L，防腐剂·························0.5g/L；2）试剂R2的组分为：缓冲液·························100mmol/L，腺苷·························...
【专利技术属性】
技术研发人员：李志明，甘宜梧，谭柏清，王珍，李贵雨，
申请(专利权)人：山东博科生物产业有限公司，
类型：发明
国别省市：山东;37
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