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首页> 外文期刊>Combinatorial Chemistry & High Throughput Screening >High Throughput Heme Assay by Detection of Chemiluminescence of Reconstituted Horseradish Peroxidase
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High Throughput Heme Assay by Detection of Chemiluminescence of Reconstituted Horseradish Peroxidase

机译：通过检测重组辣根过氧化物酶化学发光的高通量血红素测定

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In living organisms, heme is an essential molecule for various biological functions. Recent studies also suggest that heme functions as organelle-derived signal that regulates fundamental cell processes. Furthermore, estimation of heme is widely used for studying various blood disorders. In this regard, development of a rapid, sensitive, and high throughput heme assay has been sought. The most frequently used method of measuring heme by pyridine hemochrome is time, labor, and material intensive, and therefore limiting in its utility for large scale, high throughput analysis. Recently, we reported alternative method that is sensitive and specific to heme, which is based on the ability of horseradish peroxidase (HRP) apo-enzyme to reconstitute with heme to form an active holo-enzyme. Here, we developed high throughput heme assay by performing reactions on multi-well plate with highly sensitive chemiluminescence detection reagents. Detection of chemiluminescence in charged coupled device (CCD)-based gel doc apparatus enables simultaneous measurement of multiple samples. Furthermore, the high sensitivity of this assay allowed a direct measurement of heme in solvent extracts after dilution. This assay is sensitive, quick, provides a large dynamic range, and is well suited for large-scale analysis of heme extracted from minute amount of samples.
机译：在活生物体中，血红素是各种生物学功能所必需的分子。最近的研究还表明，血红素起细胞器来源的信号的作用，调节基本的细胞过程。此外，血红素的估计被广泛用于研究各种血液疾病。在这方面，已经寻求开发快速，灵敏和高通量的血红素测定。通过吡啶血色素测量血红素的最常用方法是时间，劳力和材料密集型，因此限制了其在大规模，高通量分析中的实用性。最近，我们报道了对血红素敏感且特异的另一种方法，该方法基于辣根过氧化物酶（HRP）脱辅酶与血红素重构形成活性全酶的能力。在这里，我们通过在多孔板上与高灵敏度化学发光检测试剂进行反应，开发了高通量血红素测定法。在基于电荷耦合器件（CCD）的凝胶doc仪器中检测化学发光可同时测量多个样品。此外，该测定法的高灵敏度允许稀释后直接测量溶剂提取物中的血红素。该测定灵敏，快速，提供大的动态范围，非常适合大规模分析从微量样品中提取的血红素。

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