Tomographic Phase Microscopy (TPM) - Biomedical Optical Phase Microscopy and Nanoscopy - page 243

compared with the rest of the cell suggests that increased whitening of precancerous cells
may reflect the greater nuclear-to-cytoplasmic volume ratio in such cells [32] .

A similar analysis is performed to monitor changes in shape and structure of a single cell
during exposure to a hypertonic-buffered saline solution. Figure 12.13 shows an HT29
(human colonic adenocarcinoma) cell, during a change in solution osmolarity from 300 mosm/l
to 975 mosm/l. To determine the changes in index of refraction of different components of the
cell, a 2D mask is drawn around a section of the cytoplasm, nucleolus, and nucleus regions.
Because the boundary of the cytoplasm and other organelles vary slightly over the course of the
video, the masks are drawn to maintain validity throughout the video. The average index of
refraction inside each mask is calculated over the 15.4 s of recording time. After exposure to
the hyperosmolar solution, the cell shrinks, and the average nuclear and cytoplasmic refractive
indices exhibit a roughly linear increase of approximately 1.6 3 10 2 3 /s and 1.7 3 10 2 3 /s,
respectively. The nucleolar refractive index increases only slightly.

In summary of this section, the use of spatial fringe pattern demodulation enables the
acquisition of tomograms about 300 times faster than with the original TPM based on
phase-shifting technique. The improved system is used to measure region-specific temporal
dynamics of refractive index upon changes in acidity and osmolarity. Video-rate acquisition
will also make it possible to acquire tomograms of flowing cells, with applications to
studies of cell structure using flow cytometry or microfluidic chambers [33] .

τ = 0 s τ = 7.6 s

(A) (B)

(D)

1.450

1.440

1.430

1

1.420

τ = 15.4 s

(C) 1.44

3

1.410

1.42

1.400

2

1.4

1.390

1.38

0 2 4 6 8 10 12 14 16

Time (s)

1.36

1.34

Figure 12.13
Refractive index tomograms (xy slice) of an HT29 cell during exposure to hyperosmolar
solution at time (A) 0.0 s, (B) 7.6 s, and (C) 15.4 s (movie available in Supplemental Movie S2).
(D) Time-dependent average refractive index of three ROIs as described in text [25] .