Automated method for subtraction of fluorescence from biological Raman spectra
Automated method for subtraction of fluorescence from biological Raman spectra
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DOI：
10.1366/000370203322554518
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发表时间：
2003-11-01
影响因子：
3.5
通讯作者：
Mahadevan-Jansen, A
中科院分区：
文献类型：
--
作者：
Lieber, CA;Mahadevan-Jansen, A
将拉曼光谱用于生物应用的挑战之一是测量光谱背后的许多生物分子产生的固有荧光。这种荧光有时可能比弱拉曼散射强几个数量级，并且必须尽量减少其存在，以便解析和分析拉曼光谱。为此目的，已经设计了几种涉及硬件和软件的技术；其中包括使用波长移位、时间选通、频域滤波、一阶和二阶导数以及使用高阶多项式对宽带变化进行简单曲线拟合。其中，多项式拟合被发现是一种简单但有效的方法。然而，这种技术通常需要用户干预，因此耗时且容易发生变化。描述了一种基于对最小二乘多项式曲线拟合的修改的荧光扣除的自动化方法。结果表明，所提出的自动化方法在荧光减法、重复性和拉曼光谱线形的保留方面非常出色。
One of the challenges of using Raman spectroscopy for biological applications is the inherent fluorescence generated by many biological molecules that underlies the measured spectra. This fluorescence can sometimes be several orders of magnitude more intense than the weak Raman scatter, and its presence must be minimized in order to resolve and analyze the Raman spectrum. Several techniques involving hardware and software have been devised for this purpose; these include the use of wavelength shifting, time gating, frequency-domain filtering, first- and second-order derivatives, and simple curve fitting of the broadband variation with a high-order polynomial. Of these, polynomial fitting has been found to be a simple but effective method. However, this technique typically requires user intervention and thus is time consuming and prone to variability. An automated method for fluorescence subtraction, based on a modification to least-squares polynomial curve fitting, is described. Results indicate that the presented automated method is proficient in fluorescence subtraction, repeatability, and in retention of Raman spectral lineshapes.