Department of Biomedical Engineering and Cell & Developmental Biology, Oregon Health & Science University, 3303 S. W. Bond Avenue, Portland, Oregon 97239, USA.
Phys Rev Lett. 2012 Sep 14;109(11):118105. doi: 10.1103/PhysRevLett.109.118105. Epub 2012 Sep 13.
Phase contrast microscopy has become ubiquitous in the field of biology, particularly in qualitative investigations of cellular morphology. However, the use of quantitative phase retrieval methods and their connection to cellular refractive index and dry mass density remain under utilized. This is due in part to the restriction of phase and cellular mass determination to custom built instruments, involved mathematical analysis, and prohibitive sample perturbations. We introduce tomographic bright field imaging, an accessible optical imaging technique enabling the three dimensional measurement of cellular refractive index and dry mass density using a standard transillumination optical microscope. The validity of the technique is demonstrated on polystyrene spheres. The technique is then applied to the measurement of the refractive index, dry mass, volume, and density of red blood cells. This optical technique enables a simple and robust means to perform quantitative investigations of engineered and biological specimens in three dimensions using standard optical microscopes.
相差显微镜在生物学领域已经无处不在，特别是在细胞形态的定性研究中。然而，定量相位恢复方法的使用及其与细胞折射率和干物质密度的联系仍然没有得到充分利用。这部分是由于相位和细胞质量的确定受到限制，只能在定制的仪器上进行，需要复杂的数学分析，并且对样本有干扰。我们引入了层析明场成像技术，这是一种易于使用的光学成像技术，能够使用标准的透射光显微镜对细胞折射率和干物质密度进行三维测量。该技术在聚苯乙烯球上得到了验证。然后，该技术被应用于红细胞折射率、干质量、体积和密度的测量。这种光学技术为使用标准光学显微镜对工程和生物样本进行三维定量研究提供了一种简单而强大的手段。
