Rapid detection of methylglyoxal in blood samples

快速检测血液样本中的甲基乙二醛

• 批准号: 8455478
• 负责人: JOSEPH FRANCIS KREBS
• 金额: $ 16.17万
• 依托单位: BIOO SCIENTIFIC CORPORATION
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-01-22 至 2016-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8455478
• 关键词: Affect; American; Biological Assay; Biological Factors; Biological Markers; Blood; Blood Circulation; Blood Proteins; Blood Vessels; Blood specimen; Buffers; Cardiovascular Diseases; Cause of Death; Clinical; Complications of Diabetes Mellitus; Country; Coupled; Detection; Diabetes Mellitus; Diagnostic; Disease; Dyes; Enzymes; Equilibrium; Glucose; Goals; Health; Heart Diseases; High Pressure Liquid Chromatography; Housing; Hypertension; Incidence; Investigation; Lead; Life; Link; Measures; Mediating; Methods; Monitor; Mus; NADH; NADP; Neuropathy; Oxidoreductase; Pathogenesis; Phase; Plasma; Play; Preclinical Testing; Proteins; Pyruvaldehyde; Rattus; Reaction; Reader; Reagent; Recombinants; Research; Research Personnel; Rodent; Role; Sampling; Scientist; Serum; Signal Transduction; Testing; Time; Validation; absorption; analytical method; base; cardiovascular injury; combat; diabetic; glucose metabolism; microbial; novel; novel diagnostics; oxidation; pandemic disease; pre-clinical; prevent; public health relevance; rapid detection; sugar; tool

DESCRIPTION (provided by applicant): Diabetes is a serious, life-threatening disease caused by improper absorption and metabolism of glucose. Increased glucose levels can lead to an accumulation of toxic sugar degradation products, such as methylglyoxal (MG), in the bloodstream. MG is a reactive molecule which can modify and inactivate blood proteins which in turn leads to hypertension, neuropathy, and heart disease. Consequently, MG has been shown to be an excellent biomarker for a number of diabetes-related complications. Unfortunately, all current methods to directly measure MG levels in blood are very expensive, time-consuming and laborious. To fully realize the diagnostic potential of MG, rapid analytical methods are needed for the routine determination of MG in biomedical samples such as blood. Our proposed Phase I research will create a unique test to measure MG levels in blood samples such as plasma and serum. This unique test will permit researchers to conveniently study the accumulation of reactive MG in blood before damage to proteins and vasculature has occurred. Our novel assay will use a recombinant bacterial oxidoreductase enzyme (MG-R) capable of specifically and rapidly converting MG into a detectable colorimetric signal in a 96 well plate assay format. To boost sensitivity, we will link our enzymatic detection reaction to an NADP+/NADPH visible colorimetric amplification loop to increase assay sensitivity more than 100-fold. The assay will be optimized and validated using rat and mouse plasma and serum spiked with known amounts of MG. After validation, we will commercially launch a rapid test kit to directly detect MG in preclinical and other biomedical samples.

描述（由申请人提供）：糖尿病是一种严重的、危及生命的疾病，由葡萄糖的吸收和代谢不当引起。葡萄糖水平升高可导致血液中有毒糖降解产物（如甲基乙二醛（MG））的积累。MG是一种反应性分子，可以改变和抑制血液蛋白，这反过来又导致高血压、神经病和心脏病。因此，MG已被证明是许多糖尿病相关并发症的极好生物标志物。不幸的是，目前所有直接测量血液中MG水平的方法都非常昂贵，耗时且费力。为了充分实现MG的诊断潜力，需要快速分析方法用于常规测定生物医学样品（如血液）中的MG。我们提出的I期研究将创建一个独特的测试来测量血液样本中的MG水平，如血浆和血清。这种独特的测试将使研究人员能够方便地研究在蛋白质和血管损伤发生之前，反应性MG在血液中的积累。我们的新测定将使用重组细菌氧化还原酶（MG-R），其能够在96孔板测定格式中特异性地且快速地将MG转化为可检测的比色信号。为了提高灵敏度，我们将我们的酶检测反应连接到NADP+/NADPH可见光比色放大环，以提高检测灵敏度超过100倍。将使用加标已知量MG的大鼠和小鼠血浆和血清对试验进行优化和验证。经过验证后，我们将商业化推出快速检测试剂盒，直接检测临床前和其他生物医学样本中的MG。