CN110045100B - 一种慢性阻塞性肺疾病分型试剂盒 - Google Patents
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Abstract
本发明涉及体外诊断领域，尤其涉及一种慢性阻塞性肺疾病分型的标志物。本发明提供了检测血清中的Immunoglobulin heavy variable 3/OR16‑9(non‑functional)蛋白的试剂在制备慢性阻塞性肺疾病分型试剂盒中的用途；所述分型指：区分慢性支气管炎型或肺气肿型慢性阻塞性肺疾病。本发明可借助检测Immunoglobulin heavy variable 3/OR16‑9(non‑functional)蛋白在血清中的含量，实现慢性阻塞性肺疾病的有效分型。
Description
技术领域
本发明涉及体外诊断领域，尤其涉及一种慢性阻塞性肺疾病分型的试剂盒。
背景技术
慢性阻塞性肺疾病(chronic obstructive pulmonary disease，COPD)是一种具有气流阻塞特征的慢性支气管炎和(或)肺气肿，可进一步发展为肺心病和呼吸衰竭的常见慢性疾病。与有害气体及有害颗粒的异常炎症反应有关，致残率和病死率很高，全球40岁以上发病率已高达9％～10％。
COPD的确切病因不清楚，一般认为与慢性支气管炎和阻塞性肺气肿发生有关的因素都可能参与慢性阻塞性肺病的发病。对COPD进行正确的分型——慢支炎(慢性支气管炎)型或肺气肿型，可为COPD治疗提供有价值的参考。
发明内容
本发明的目的在于提供一种COPD分型标志物和试剂盒，所述分型指区分慢性支气管炎型或肺气肿型慢性阻塞性肺疾病。
本发明所述的Immunoglobulin heavy variable 3/OR16-9(non-functional)蛋白，指的是UniProt知识库中的编号为A0A0B4J2B5的蛋白。
技术方案如下：
检测血清中的Immunoglobulin heavy variable 3/OR16-9(non-functional)蛋白的试剂在制备慢性阻塞性肺疾病分型试剂盒中的用途；所述分型指：区分慢性支气管炎型或肺气肿型慢性阻塞性肺疾病。
如前所述的用途，所述检测血清中的Immunoglobulin heavy variable 3/OR16-9(non-functional)蛋白的试剂为：免疫组化检测方法用试剂、Western Blot检测方法用试剂、ELISA检测方法用试剂、TMT定量蛋白质组学方法用试剂或R&D Systems Luminex液相芯片检测用试剂。
如前所述的用途，所述试剂还包括蛋白纯化试剂。
一种慢性阻塞性肺疾病分型检测试剂盒，所述试剂盒包括检测Immunoglobulinheavy variable 3/OR16-9(non-functional)蛋白的试剂。
如前述的试剂盒，所述检测Immunoglobulin heavy variable 3/OR16-9(non-functional)蛋白的试剂为免疫组化检测方法用试剂。
如前述的试剂盒，所述检测Immunoglobulin heavy variable 3/OR16-9(non-functional)蛋白的试剂为Western Blot检测方法用试剂。
如前述的试剂盒，所述检测Immunoglobulin heavy variable 3/OR16-9(non-functional)蛋白的试剂为ELISA检测方法用试剂。
如前述的试剂盒，所述检测Immunoglobulin heavy variable 3/OR16-9(non-functional)蛋白的试剂为TMT定量蛋白质组学方法用试剂。
如前述的试剂盒，所述检测Immunoglobulin heavy variable 3/OR16-9(non-functional)蛋白的试剂为R&D Systems Luminex液相芯片检测用试剂。
如前述的任一试剂盒，它还包括蛋白纯化试剂。
Immunoglobulin heavy variable 3/OR16-9(non-functional)蛋白在慢支炎型COPD患者的血清中显著低于肺气肿型COPD患者，通过检测COPD患者血清中Immunoglobulinheavy variable 3/OR16-9(non-functional)蛋白能够有效对COPD进行分型。
本发明提供的标志物可以被多种蛋白检测方法检测得到，可开发成多种COPD分型试剂盒。
本发明的试剂盒能够提供快速准确地对COPD进行分型，为医生提供诊断依据，具有良好的市场前景。
显然，根据本发明的上述内容，按照本领域的普通技术知识和惯用手段，在不脱离本发明上述基本技术思想前提下，还可以做出其它多种形式的修改、替换或变更。
以下通过具体实施方式对本发明的上述内容再作进一步的详细说明。但不应将此理解为本发明上述主题的范围仅限于以下的实例。凡基于本发明上述内容所实现的技术均属于本发明的范围。
具体实施方式
实施例1不同表型COPD(慢支炎型/肺气肿型)中Immunoglobulin heavy variable3/OR16-9(non-functional)蛋白的检测
1.被检对象
慢支炎型COPD患者(A组)30人，肺气肿型COPD患者(B组)30人。被检对象人群在实验前均已经知情并同意。
2.方法
取被检对象的血清提取蛋白，进行TMT定量蛋白质组学检测，具体地，包括如下步骤：
将慢支炎表型与肺气肿表型COPD患者血清各取5个样，使用不同TMT试剂进行标记，等量混合。离心后取上清液，去除高丰度蛋白，用BCA试剂盒进行蛋白浓度测定，洗脱液分别取20μg进行SDS-PAGE电泳检测高丰度蛋白去除情况。胰酶酶解肽段用Strata X C18(Phenomenex)除盐后真空冷冻干燥。以0.5M TEAB溶解肽段，肽段采用高pH反向HPLC分级。肽段采用EASY-nLC 1000超高效液相系统分离，并同步使用Thermo ScientificTMQExactiveTMPlus质谱系统进行检测和分析。肽段分离后进行电离，然后进入Q ExactiveTMPlus质谱进行分析。肽段母离子及其二级碎片均采用Orbitrap检测和分析。二级质谱数据采用Mascot search engine(v.2.3.0)检索，同时进行质谱质控。
该方法每次能检测10个样品，以此方法分批检测剩余血清样，得到60名被检对象的血清中的蛋白质组学信息，其中包括Immunoglobulin heavy variable 3/OR16-9(non-functional)蛋白的含量信息。
3.结果
A、B组血清中的Immunoglobulin heavy variable 3/OR16-9(non-functional)蛋白检测情况如下表所示：
| A组均值 | B组均值 | FC | PVALUE |
| 0.670049 | 2.168358 | 0.309012132 | 5.07534E-05 |
备注：FC(fold change)是指的该蛋白在慢支炎表型血清中的含量/肺气肿表型血清中的含量
可见，慢支炎型COPD患者血清中的Immunoglobulin heavy variable 3/ORl6-9(non-functional)蛋白的含量显著低于肺气肿型COPD患者。基于Immunoglobulin heavyvariable 3/OR16-9(non-functional)蛋白在COPD患者血清中的含量的检测，可开发出试剂盒，实现慢支炎型和肺气肿型COPD的有效分型。
实施例2本发明的试剂盒
1.本发明的试剂盒的组成：
| TMT试剂 |
| 尿素 |
| 硫脲 |
| 二硫苏糖醇 |
| 碘乙酰胺 |
| 甲酸 |
| 蛋白酶抑制剂 |
2.试剂盒使用方法
试剂盒的使用方法包括如下步骤：
蛋白提取—BCA蛋白浓度测定-SDS-聚丙烯酰胺凝胶电泳-胰蛋白酶酶解及标记-反相色谱分离。
具体地，同实施例1的第2节。
本发明的试剂盒通过对血清中Immunoglobulin heavy variable 3/OR16-9(non-functional)蛋白的检测，可实现对慢支炎型或肺气肿型COPD的有效分型检测。
Claims (3)
1.检测血清中的Immunoglobulin heavy variable 3/OR16-9(non-functional)蛋白的试剂在制备慢性阻塞性肺疾病分型试剂盒中的用途；所述分型指：区分慢性支气管炎型或肺气肿型慢性阻塞性肺疾病；所述Immunoglobulin heavy variable 3/OR16-9(non-functional)蛋白为UniProt知识库中的编号为A0A0B4J2B5的蛋白。
2.如权利要求1所述的用途，其特征在于，所述检测血清中的Immunoglobulinheavyvariable 3/OR16-9(non-functional)蛋白的试剂为：免疫组化检测方法用试剂、WesternBlot检测方法用试剂、ELISA检测方法用试剂、TMT定量蛋白质组学方法用试剂或R&DSystems Luminex液相芯片检测用试剂。
3.如权利要求2所述的用途，其特征在于，所述试剂还包括蛋白纯化试剂。
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| CN108303547A (zh) * | 2018-02-07 | 2018-07-20 | 北京泱深生物信息技术有限公司 | 一种用于诊断慢性阻塞性肺疾病的分子标志物 |
| CN109045233A (zh) * | 2018-09-30 | 2018-12-21 | 宁夏医科大学 | 用于治疗慢性阻塞性肺疾病的中药组合物、制备方法以及其应用 |
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| CN107267662A (zh) * | 2017-08-22 | 2017-10-20 | 常州市第二人民医院 | 慢性阻塞性肺疾病的诊断工具 |
| CN108303547A (zh) * | 2018-02-07 | 2018-07-20 | 北京泱深生物信息技术有限公司 | 一种用于诊断慢性阻塞性肺疾病的分子标志物 |
| CN109045233A (zh) * | 2018-09-30 | 2018-12-21 | 宁夏医科大学 | 用于治疗慢性阻塞性肺疾病的中药组合物、制备方法以及其应用 |
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| Vascular endothelial growth factor: an angiogenic factor reflecting airway inflammation in healthy smokers and in patients with bronchitis type of chronic obstructive pulmonary disease?;Nikoletta Rovina等;《Respiratory Research》;20070715;第1-8页 * |
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