CN106568944B - 一种检测AGEs的胶乳免疫比浊试剂盒 - Google Patents
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Abstract
本发明涉及胶乳免疫比浊法用于测定人血清中AGEs的试剂，具体提供的试剂包括试剂R1和试剂R2，其中试剂R1中含有碳酸盐缓冲液、氯化钠、蔗糖、牛血清白蛋白、ProClin300；试剂R2中含有碳酸盐缓冲液、氯化钠、蔗糖、牛血清白蛋白、抗人AGEs抗体的胶乳微球、聚乙二醇6000、月桂酸钾、明胶、甘油、ProClin300。该试剂稳定性和线性好，特异性强，分析灵敏度高，操作简便，易于保存等优点，可用于临床中常用的全自动生化分析仪。
Description
技术领域
本发明涉及一种体外诊断试剂领域，具体涉及一种AGEs检测试剂，还涉及使用此检测试剂的检测方法。
背景技术
AGEs是以蛋白质、脂肪及核酸的氨基和还原糖发生非酶催化反应产生稳定的、不可逆的化合物，同时它能够和人体的组织和细胞相结合并造成严重的破坏。它在生物体中的来源，一方面多量的糖会和蛋白质发生此反应，另一方面进食的食物中存在该物质被摄入体内。由于它的积聚会引发糖尿病等各种并发症，如糖尿病肾病、阿尔兹海默症、动脉粥样硬化、白内障等症，影响全身各系统，因此对其进行快捷、简单并准确的检验在临床诊断中具有重要意义。目前它的检测方法有放射受体分析法、放射免疫法、酶联免疫吸附法、高效液相色谱法等，但是这些方法操作过程繁琐，误差大，消耗时间长，自动化程度低，费用高，灵敏度低，对于临床非常不利，一般只用来定性检测。
流行病学和临床研究表明，AGEs与多种疾病的密切相关，现今对其治病机制研究已经取得了很大的进展，同时相关靶点的药物也在不断开发，所以对其进行精确的诊断成为了重要先导步骤。
发明内容
本发明的目的是提供一种用于检测AGEs的试剂以及使用该试剂检测AGEs含量的方法。采用胶乳免疫比浊法的方式检测血清中AGEs的含量，包括游离态的AGEs和结合态的AGEs，线性好，灵敏度高，特异性好、操作简单、方便快速等优点。它能够作为一项新的临床指标，AGEs检测对阐明这些疾病的发病机制和判定治疗效果有一定价值。
本发明采用以下步骤：
一种AGEs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
试剂R2中含有
本发明的优点：
1.采用的表面活性剂为月桂酸钾和聚乙二醇6000的用量比例为2∶1，显著改善了连接抗体微球的稳定性。
2.采用的被连接胶乳微球的粒径分别为300nm，100nm，50nm，并且三种胶乳微球的所占比例为1∶3∶2，提升了检测的线性和灵敏度。
3.采用的稳定剂组合为蔗糖、明胶、甘油和牛血清白蛋白，增加了试剂的稳定性。
4.试剂的稳定性好，准确度高，使用方便，满足临床需要。
附图说明：
图1为实施例1试剂检测结果与对照方法检测结果相关分析图，采用BS-420全自动生化分析仪，对40个血清样本进行测定，并对测定值进行相关分析。其中相关系数：r2＝0.999，线性方程为：y＝1.011x-0.124。
具体实施方式
下面结合具体实施例对本发明进行进一步说明：
实施例1
AGEs检测试剂，包括试剂R1和试剂R2：
试剂R1中含有
试剂R2中含有
本实施例描述的AGEs检测试剂，使用双试剂全自动生化分析仪，例如贝克曼DXC800、日立7600等全自动分析仪，选用终点法进行测定。将R1和R2放置所设置好的位置上，在样本盘中对应位置放好蒸馏水、标准品和待测样本，操作如下：
表1操作程序
结果计算：以测定管ΔA，根据校准曲线可求得AGEs含量。
实施例2
相关性试验：利用实施例1中试剂，与荧光光谱法作对比，同时检测了40例临床血清样本，检测结果表2所示，结果证明二者具有很好的相关性。
表2实施例1试剂与荧光光谱法检测AGEs含量的方法对比检测结果(ng/mL)
实施例3
本实例只改变胶乳微球的粒径大小和所占比重，选用同一个样本检测，实施例3与实施例1中线性关系如表3所示，选取标准品浓度为30ng/mL，结果显示实施例1试剂的线性较实施例3好。
表3线性关系结果
| 序号 | 实施例1试剂 | 实施例3试剂 |
| 0 | 0 | 0 |
| 1 | 3.6 | 3.74 |
| 2 | 10.2 | 7.51 |
| 3 | 18.9 | 15.2 |
| 4 | 29.3 | 29.3 |
| 相关系数 | 0.996 | 0.929 |
分析灵敏度验证试验：
用实施例1、实施例3和荧光光谱法(对比方法)检测已知浓度的样本，记录吸光度差值。检测结果如表4所示。
表4分析灵敏度试验结果
综合上述结果可知，实施例1中胶乳微球粒径大小和所占的比例，该试剂有较好的灵敏度和线性。
Claims (1)
1.一种AGEs检测试剂，其特征包括试剂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
试剂R2中含有
；
所用的抗人AGEs抗体胶乳微球的粒径分别为300nm，100nm，50nm，并且三种胶乳微球的用量质量比例为1∶3∶2。
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