## 3 Cell Death Studies with QPI  
The ability of QPI to differentiate between viable and non-viable cells without the use of staining has been extensively studied [21; 83]. The first DH microscopy images of tumor spheroids were published in 2003 [84], and the first images of single cells one year after that [85]. Indeed, DH microscopy has turned out to be a very useful technique for cell death studies [21; 83]. Cells that go into programmed cell death, or apoptosis, show a decreased cell phase shift as the first indication of early apoptosis, but several morphological alterations take place, such as cell shrinkage, formation of small blebs, nuclear fragmentation, chromatin condensation, chromosomal DNA fragmentation, and breakdown into several apoptotic bodies [86]. [PERSON] et al. showed that dead cells could be identified within minutes through their DH phase signal, whereas with conventional trypan blue staining the identification took several hours [87]. Also, DH microscopy was compared with electronic cell sizing and atomic force microscopy [88]. DH was shown to be advantageous in terms of the noninvasive labeling, the time resolution, and the possibility of measuring both single cells and cell populations. Moreover, in a study by [PERSON] et al., DH measurements revealed significant differences in the average cell number, the confluence, cell volume, and cell area when comparing untreated and etoposide-treated cells [50]. We have cultured T leukemia Jurkat cells in ibidi chambers (ibidi GmbH, Martinsried, Germany) and treated the cells for 24 h with the cell death-inducing agent etoposide, or left untreated as a negative control. After the incubation, the cells were analyzed with DH microscopy. 3D holograms are shown for untreated cells (Figure 2A) and for the etoposide-treated cells (Figure 2B). Interestingly, several DH parameters that were measured, including cell number, area, thickness, and volume, were decreased after Etposide treatment of the Jurkat cells (Table 2). \begin{table}
\begin{tabular}{c c c c c c} \hline \hline  & **Number of** & **Confluence** & **Av. Cell** & **Av. Cell Optical** & **Av. Cell Optical** & **Volume (\(\upmu\)m\({}^{3}\))** \\ \hline Control & 226 \(\pm\) 81 & 10.9 \(\pm\) 3.8 & 143.4 \(\pm\) 7.0 & 6.5 \(\pm\) 0.4 & 1006.2 \(\pm\) 121.1 \\ Etposide & 114 \(\pm\) 17 & 4.4 \(\pm\) 0.5 & 115.0 \(\pm\) 8.4 & 4.9 \(\pm\) 0.2 & 584.0 \(\pm\) 48.2 \\ \hline \hline \end{tabular}
\end{table} 
Table 2: Quantitative DH measurements of Jurkat cells treated with 250 \(\upmu\)M etoposide for 24 h.  
Figure 2: Jurkat cells cultured in ibidi chambers for 24 h: (**A**) Untreated cells; (**B**) Cells treated with 250 \(\upmu\)M etoposide. [PERSON] et al. have shown details of the apoptotic process, where the apoptotic blebbing in prostate cancer cells was clearly visualized by DH microscopy [81]. Interestingly, results presented on the individual cellular level of prostate cancer cells and mouse fibroblast cells treated with cell death-inducing drugs showed that these cell types behaved very differently [82]. Using rapid profiling with high-speed live-cell interferometry, [PERSON] and co-workers recently showed that the proto-oncogene B-Raf (BRAF)-inhibitor sensitive melanoma cell lines differed in measurements of optical cell biomass [62]. Interestingly, within 24 h, biomass kinetic signatures were obtained for three pairs of sensitive or resistant melanoma cell lines, which showed a discrimination. Cell death and cell proliferation are stringently controlled by the cell cycle, and studies have shown the ability of QPI to identify the different phases of the cell life cycle, i.e., specific changes in cell phase correlating to either a G1 or a G2/M arrest [13; 25; 38; 44; 57; 89]. [PERSON] et al. constructed individual cell trees showing the way in which the drug salinomycin affects individual cells cultured in normoxia or hypoxia, by monitoring the times to first and last division [13]. In this way, sensitivity to different conditions could be analyzed.