Title: Refractive index measurements of single, spherical cells using digital holographic microscopy,
Abstract: In this chapter, we introduce digital holographic microscopy (DHM) as a
marker-free method to determine the refractive index of single, spherical cells
in suspension. The refractive index is a conclusive measure in a biological
context. Cell conditions, such as differentiation or infection, are known to
yield significant changes in the refractive index. Furthermore, the refractive
index of biological tissue determines the way it interacts with light. Besides
the biological relevance of this interaction in the retina, a lot of methods
used in biology, including microscopy, rely on light-tissue or light-cell
interactions. Hence, determining the refractive index of cells using DHM is
valuable in many biological applications. This chapter covers the main topics
which are important for the implementation of DHM: setup, sample preparation
and analysis. First, the optical setup is described in detail including notes
and suggestions for the implementation. Following that, a protocol for the
sample and measurement preparation is explained. In the analysis section, an
algorithm for the determination of the quantitative phase map is described.
Subsequently, all intermediate steps for the calculation of the refractive
index of suspended cells are presented, exploiting their spherical shape. In
the last section, a discussion of possible extensions to the setup, further
measurement configurations and additional analysis methods are given.
Throughout this chapter, we describe a simple, robust, and thus easily
reproducible implementation of DHM. The different possibilities for extensions
show the diverse fields of application for this technique.