NAD(P)H fluorescence lifetime measurements in fixed biological tissues
NAD(P)H fluorescence lifetime measurements in fixed biological tissues
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DOI：
10.1088/2050-6120/ab47e5
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发表时间：
2019-10-01
影响因子：
3.2
通讯作者：
Eliceiri, Kevin W.
中科院分区：
文献类型：
--
作者：
Chacko, Jenu V.;Eliceiri, Kevin W.
基于自体荧光的荧光寿命成像显微镜（AF-FLIM）技术已经从癌症表征的早期研究中走了很长的路，现在已经广泛应用于涵盖广泛疾病的几种细胞和动物研究中。自体荧光成像（AFI）的大多数研究都是研究活体生物样品中的代谢通量。然而，来自临床或科学研究的组织通常被化学固定以保存和稳定组织形态。固定对于酶、功能或组织病理学研究尤其重要。由于缺乏该领域的研究，代谢成像（如光学氧化还原强度成像和AF-FLIM）的解释通常被视为在固定样本中可能不可靠。在这项研究中，我们仔细评估的可能性，提取微环境的信息，在固定的组织使用减少烟酰胺腺嘌呤二核苷酸（磷酸）（NAD（P）H）内源性荧光。在固定的组织中使用固有荧光区分诸如代谢和pH的变化的能力具有很大的病理学价值。在这项工作中，我们表明，在化学固定后，保存在样品中的基于寿命的代谢对比度。样品的荧光寿命随着添加剂固定剂如甲醛而增加;然而，固定的组织即使在固定后也保留代谢特征。本研究提供了一个机会，成功地成像存档的未染色的组织病理学组织，并生成有用的AF-FLIM签名。我们证明了即使在长时间储存后，在固定的组织中绘制代谢解释的能力。
Autofluorescence based fluorescence lifetime imaging microscopy (AF-FLIM) techniques have come a long way from early studies on cancer characterization and have now been widely employed in several cellular and animal studies covering a wide range of diseases. The majority of research in autofluorescence imaging (AFI) study metabolic fluxes in live biological samples. However, tissues from clinical or scientific studies are often chemically fixed for preservation and stabilization of tissue morphology. Fixation is particularly crucial for enzymatic, functional, or histopathology studies. Interpretations of metabolic imaging such as optical redox intensity imaging and AF-FLIM, have often been viewed as potentially unreliable in a fixed sample due to lack of studies in this field. In this study, we carefully evaluate the possibility of extracting microenvironment information in fixed tissues using reduced nicotinamide adenine dinucleotide (phosphate) (NAD(P)H) endogenous fluorescence. The ability to distinguish changes such as metabolism and pH using intrinsic fluorescence in fixed tissues has great pathological value. In this work, we show that the lifetime based metabolic contrast in a sample is preserved after chemical fixation. The fluorescence lifetime of a sample increases with an additive fixative like formaldehyde; however, the fixed tissues retain metabolic signatures even after fixation. This study presents an opportunity to successfully image archived unstained histopathology tissues, and generate useful AF-FLIM signatures. We demonstrate the capability to draw metabolic interpretations in fixed tissues even after long periods of storage.