Multispectral fluorescence imaging to assess pH in biological specimens
Multispectral fluorescence imaging to assess pH in biological specimens
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DOI：
10.1117/1.3533264
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发表时间：
2011-01-01
影响因子：
3.5
通讯作者：
Manning, H. Charles
中科院分区：
文献类型：
--
作者：
Hight, Matthew R.;Nolting, Donald D.;Manning, H. Charles
测量组织中pH值的简单定量分析可以改善对复杂生物过程和癌症等疾病的研究。我们评估了多光谱荧光成像（MSFI），以定量染料灌注，切除肿瘤标本与市售仪器的细胞外pH值（pH（e））。利用具有pH依赖性荧光发射的水溶性有机染料（SNARF-4F），我们使用标准荧光法定量评估染料的发射特性作为pH值的函数。通过在成像系统提供的适当成像滤光片组施加的光谱限制内进行这些研究，我们确定，由于次优激发，去质子化染料的荧光发射的校正对于精确测定pH是必要的。随后，采用荧光衍生的校正因子（C-F），MSFI数据集的染料水溶液和组织样phantomy可以光谱解混，以准确地量化质子化（HA）和去质子化（A（-））染料的平衡浓度，从而确定溶液pH值。最后，我们探讨了MSFI的可行性高分辨率pH值（e）映射的人结直肠癌细胞系异种移植物。提供的数据表明，MSFI适用于定量测定离体染料灌注组织中的pH（e），可能能够测量各种临床前疾病模型中的pH。(C)2011年，美国光电仪器工程师学会（SPIE）。[DOI：10.1117/1.3533264]
Simple, quantitative assays to measure pH in tissue could improve the study of complicated biological processes and diseases such as cancer. We evaluated multispectral fluorescence imaging (MSFI) to quantify extracellular pH (pH(e)) in dye-perfused, surgically-resected tumor specimens with commercially available instrumentation. Utilizing a water-soluble organic dye with pH-dependent fluorescence emission (SNARF-4F), we used standard fluorimetry to quantitatively assess the emission properties of the dye as a function of pH. By conducting these studies within the spectroscopic constraints imposed by the appropriate imaging filter set supplied with the imaging system, we determined that correction of the fluorescence emission of deprotonated dye was necessary for accurate determination of pH due to suboptimal excitation. Subsequently, employing a fluorimetry-derived correction factor (C-F), MSFI data sets of aqueous dye solutions and tissuelike phantoms could be spectrally unmixed to accurately quantify equilibrium concentrations of protonated (HA) and deprotonated (A(-)) dye and thus determine solution pH. Finally, we explored the feasibility of MSFI for high-resolution pH(e) mapping of human colorectal cancer cell-line xenografts. Data presented suggest that MSFI is suitable for quantitative determination of pH(e) in ex vivo dye-perfused tissue, potentially enabling measurement of pH across a variety of preclinical models of disease. (C) 2011 society of Photo-Optical Instrumentation Engineers (SPIE). [DOI: 10.1117/1.3533264]