Rapid diagnosis of the systemic inflammatory response. syndrome by the transcription factor activation measuring method development concerning the inflammation

快速诊断全身炎症反应。

基本信息

批准号：
17590486
负责人：
KITAJIMA Isao
金额：
$ 2.24万
依托单位：
University of Toyama
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2006
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-17590486/
关键词：
systemic inflammatory response syndrome (SIRS)inflammatory cytokine rapid testing transcriptional factor Nuclear factor-kappa B sepsis ELISA surface plasmon resonance (SPR)1分子計測

项目摘要

The nuclear factor-kappa B (NF-κB) family of transcription factors is known to play an important role in the regulation of the immune system. NF-κB is activated by bacterial and viral antigens, which leads to the production of proinflammatory cytokines and chemokines. The rapid detection of activated NF-κB activation by the systemic inflammatory response syndrome (SIRS) is considered to be crucial for the treatment of patients with septicemia. The aim of the present study was to evaluate the sensitivity of two methods, electrophoretic mobility shift assay (EMSA) and transcription factor enzyme-linked immunoassay (TF-ELISA). TF-ELISA detected 25 ng of recombinant human NF-κB p50 (rhp50) and 5 mg of TNFα-stimulated HeLa nuclear protein, while EMSA detected approximately 100 ng of rhp50 and 10 mg of HeLa nuclear protein. We found that the TF-ELISA was more sensitive than EMSA in detecting NF-κB. However, it was judged that the 3-6 hour measuring time required in TF-ELISA was excessively long for patients with SIRS. Therefore, the development of new analytical methods with improved sensitivity and, measurement time was necessary for the detection and quantification of activated NF-κB protein in the hospital laboratory. Consequently, we have developed a new NF-κB analyzer based on surface plasmon resonance (SPR), which is recognized as one of the most sensitive direct optical detection methods. This method can detect nanomolar concentrations of NF-κB within 15 minutes. At present, we are carrying out clinical trials using this new transcription factor analysis apparatus for SIRS patients.

众所周知，核因子 kappa B (NF-κB) 转录因子家族在免疫系统的调节中发挥着重要作用，NF-κB 被细菌和病毒抗原激活，从而导致促炎细胞因子的产生。全身炎症反应综合征（SIRS）对活化的 NF-κB 的快速检测被认为对于败血症患者的治疗至关重要。本研究的目的是评估两种方法的敏感性。迁移率变动测定 (EMSA) 和转录因子酶联免疫测定 (TF-ELISA) 检测到 25 ng 重组人 NF-κB p50 (rhp50) 和 5 mg TNFα 刺激的 HeLa 核蛋白，而 EMSA 检测到约100 ng rhp50 和 10 mg HeLa 核蛋白我们发现 TF-ELISA 的检测比 EMSA 更灵敏。然而，对于SIRS患者来说，TF-ELISA所需的3-6小时的测量时间被认为过长，因此，需要开发具有更高灵敏度和测量时间的新分析方法来进行检测。并在医院实验室测试活化的 NF-κB 蛋白的定量，我们开发了一种基于表面等离子共振 (SPR) 的新型 NF-κB 分析仪，该方法被认为是最灵敏的直接光学检测方法之一。 15分钟内检测纳摩尔浓度的NF-κB 目前，我们正在利用这种新型转录因子分析仪对SIRS患者进行临床试验。