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Scaffold Structure and Cell Function Through Multimodal Imaging and Quantitative Visualization

(TEMPs) are often three-dimensional (3D) hybrid materials consisting of a porous scaffold upon which the tissue is grown. While it is generally understood that a complex interaction of many variables influences the success of TEMPs, the precise nature of these interactions has yet to be worked out in many instances. A significant difficulty in furthering the understanding of the interaction between these factors and cell behavior is the lack of a high-resolution imaging technique that can penetrate deeply and nondestructively into the scaffold. An approach that uses advanced optical imaging to noninvasively monitor the developing tissue was developed. However, before any assessment of the tissue viability can be made, the volumes of imaging data must be rigorously analyzed. Therefore, an equally important component of this effort is image visualization and quantification. Progress in these areas is summarized below. MULTIMODAL IMAGING An instrument that can gather information on a TEMP using multiple imaging modalities was constructed. This means that each channel of imaging data provides different but complimentary information. Optical coherence microscopy (OCM) was chosen as the technique to image scaffold, cell, and tissue structure because of its unique combination of high resolution (ª1 µm) and high sensitivity (> 100 dB). OCM is an interferometric technique that uses both confocal and coherence gating mechanisms for stray light rejection, rendering it comparable in resolution to laser scanning confocal microscopy but far superior in imaging depth. Confocal fluorescence microscopy (CFM) was added to the OCM to collect information on cell function using traditional cell-staining techniques. In the collinear instrument, volumetric images of cell and scaffold structure were collected using the OCM channel and cell function using the CFM channel. Each channel is then overlayed in the rendered image for maximal insight. Figure 1 displays merged and registered OCM and CFM images 145 µm below the surface of a TEMP. The TEMP consists of a volume fraction of 50 percent poly(ε-caprolactone) (PCL) scaffold that was cultured with fetal chick osteoblasts for 10 weeks and stained with a nuclear stain. In Figure 1, the regions of low OCM signal are red (pores), high OCM signal are black (scaffold), and regions of high CFM signal are yellow (cells). CFM complements OCM by allowing us to positively identify stained tissues at more shallow depths. Once identified, OCM allows us to discriminate these tissues from scaffold, and thus view them at a greater depth. This will form the …