Detection of hyaluronidase activity using fluorescein labeled hyaluronic acid and Fluorescence Correlation Spectroscopy.
Detection of hyaluronidase activity using fluorescein labeled hyaluronic acid and Fluorescence Correlation Spectroscopy.
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DOI：
10.1016/j.jphotobiol.2012.07.007
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发表时间：
2012-11-05
影响因子：
5.4
通讯作者：
Fudala, Rafal
中科院分区：
文献类型：
--
作者：
Rich, Ryan M.;Mummert, Mark;Foldes-Papp, Zeno;Gryczynski, Zygmunt;Borejdo, Julian;Gryczynski, Ignacy;Fudala, Rafal
透明质酸酶的过度表达已在许多类型的癌症中观察到，这表明它可能具有诊断实用性。在这里，我们提出了一种使用荧光相关光谱（FCS）检测透明质酸酶的技术。羟丙甲环酸大分子（HA）被荧光素胺标记，导致强烈的自猝灭。在透明质酸酶的存在下，HA被切割成更小的荧光素标记的片段，并释放自猝灭。这种切割表现为HA片段的平均扩散速率增加、单个荧光HA片段的浓度增加和强度增加。在整个FCS测量过程中同时监测所有这三种性质，作为时间和透明质酸酶浓度的函数。我们提出的方法提供了透明质酸酶活性的灵敏测量，并需要极少量的HA底物。
The over-expression of hyaluronidase has been observed in many types of cancer, suggesting that it may have utility for diagnosis. Here we present a technique for the detection of hyaluronidase using Fluorescence Correlation Spectroscopy (FCS). Hyaluronan macromolecules (HAs) have been heavily labeled with fluorescein amine resulting in strong self-quenching. In the presence of hyaluronidase, HA is cleaved into smaller, fluorescein-labeled fragments and the self-quenching is released. Such cleavage is manifested by the increased average diffusion rate of the HA fragments, increased concentration of individual, fluorescent HA fragments, and increased intensity. All three of these properties are monitored simultaneously throughout FCS measurements, both as a function of time and hyaluronidase concentration. The method we present provides a sensitive measure of hyaluronidase activity and requires extremely small amounts of the HA substrate.
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DOI：
10.1016/j.jphotobiol.2011.06.003
发表时间：
2011-09-02
影响因子：
5.4
作者：
Fudala, Rafal;Mummert, Mark E.;Gryczynski, Zygmunt;Gryczynski, Lgnacy
通讯作者：
Gryczynski, Lgnacy
影响因子：
3.5
作者：
Lakowicz, JR;Gryczynski, I;Rao, G
通讯作者：
Rao, G
影响因子：
3.6
作者：
Luchowski, Rafal;Sabnis, Sushant;Gryczynski, Ignacy
通讯作者：
Gryczynski, Ignacy
影响因子：
2.9
作者：
Zhang, Li-Shu;Mummert, Mark E.
通讯作者：
Mummert, Mark E.
影响因子：
2.9
作者：
Gregor, I;Patra, D;Enderlein, J
通讯作者：
Enderlein, J