Is the nuclear refractive index lower than cytoplasm? Validation of phase measurements and implications for light scattering technologies.
Is the nuclear refractive index lower than cytoplasm? Validation of phase measurements and implications for light scattering technologies.
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DOI：
10.1002/jbio.201600314
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发表时间：
2017-12
影响因子：
2.8
通讯作者：
Wax A
中科院分区：
文献类型：
--
作者：
Steelman ZA;Eldridge WJ;Weintraub JB;Wax A
生物材料的折射率 (RI) 是生命系统光学表征的基本参数。许多光散射技术都基于对细胞和亚细胞尺度折射率的定量了解。定量相显微镜 (QPM) 的最新研究对细胞核指数大于细胞质指数这一广泛持有的假设提出了质疑，这一结果与当前的许多文献不一致。在这项工作中，我们使用 QPM 严格检查了细胞核和全细胞折射率的测量，验证了四种不同细胞系及其相应的分离细胞核中的细胞核折射率低于细胞质的折射率。我们进一步研究了米氏散射和相位包裹作为这些测量中的潜在误差源，发现它们的影响最小。最后，我们使用角度分辨散射信息通过模拟来检查不正确的 RI 假设对核形态测量的影响。尽管对核折射率的假设是错误的，但仍保持了对核形态的准确测量，这表明光散射方式仍然有效。最近使用定量相差显微镜的研究表明，核折射率低于细胞质的折射率。在本文中，我们在新细胞系（包括非癌性细胞系和原代细胞系）中验证了这一说法。我们进一步验证潜在的破坏因素，并检查低核指数对光散射技术的影响。
The refractive index (RI) of biological materials is a fundamental parameter for the optical characterization of living systems. Numerous light scattering technologies are grounded in a quantitative knowledge of the refractive index at cellular and subcellular scales. Recent work in quantitative phase microscopy (QPM) has called into question the widely held assumption that the index of the cell nucleus is greater than that of the cytoplasm, a result which disagrees with much of the current literature. In this work, we critically examine the measurement of the nuclear and whole-cell refractive index using QPM, validating that nuclear refractive index is lower than that of cytoplasm in four diverse cell lines and their corresponding isolated nuclei. We further examine Mie scattering and phase-wrapping as potential sources of error in these measurements, finding they have minimal impact. Finally, we use simulation to examine the effects of incorrect RI assumptions on nuclear morphology measurements using angle-resolved scattering information. Despite an erroneous assumption of the nuclear refractive index, accurate measurement of nuclear morphology was maintained, suggesting that light scattering modalities remain effective. Recent studies using quantitative phase microscopy have suggested that the nuclear refractive index is lower than that of cytoplasm. In this paper, we verify this claim in new cell lines, including non-cancerous and primary lines. We further validate against potential corrupting factors, and examine the impact that a low nuclear index would have on light scattering technologies.
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