The integral luminescence signal level from the labeled cells was high enough to motivate the application of UCNP in more challenging imaging scenarios, such as UCNP-assisted imaging in tissue. Since adenocarcinoma cells are hosted in human breast tissue, an imaging contrast of these cells can be modeled experimentally, provided a human breast tissue model is available. To this aim, we designed an agarose-based phantom that mimicked the optical absorption properties of live human breast tissue in the spectral ranges of the UCNP excitation and emission [Fig. 5(a)], and scattering in near-IR region. The absorption of breast tissue was calculated, considering absorption of hemoglobin (0.002 mM) and oxy-hemoglobin (0.011 mM) in the green range and near-IR light absorption of water.32 The spectrum of breast tissue absorption in the red spectral range was obtained from in vivo measurements.33 Agarose was chosen as the matrix, as its water content (about 99%) is commensurable with that of live breast tissue (10% to 60%), and resulted in slightly higher absorption of the excitation light compared to live tissue, i.e., by . The phantom and breast tissue absorption coefficients () integrated over the relevant wavelength bands were similar (green: , , red: , and 978 nm: , ), see Fig. 5(a). The reduced scattering coefficient of breast tissue in vivo was simulated by adding submicron particles to the phantom.34 The scattering coefficient and average cosine of scattering (-value) in our phantom were defined by Mie calculations of particles in water. Matching the reduced scattering coefficient at 978 nm with values recorded in vivo (978 nm: , ) resulted in a decreased scattering coefficient of the phantom compared to live tissue for the green and red wavelength bands (red: to , and green: to , ).33 Matching the scattering in live tissue and phantom at 978 nm was crucial and hence employed in our modeling, since the scattering of 978-nm light primarily determined the luminescence signal decay with the depth due to the nonlinear UCNP . Thin layers of the phantom material (0.4 to 1.4 mm) were prepared (see Sec. 2) and individually stacked between the sample plane and epi-luminescence microscope objective lens, as shown in Fig. 5(b).