Progress in bio-printing techniques has accentuated the need for non-invasive imaging modalities with high sensitivity, high resolution, and imaging depth of a few millimeters. Mesoscopic Fluorescence Molecular Tomography (MFMT) is a promising imaging modality for 3D localization and quantification of fluorescent labeled cells in thick scaffolds. Here we report on the characterization of our second generation MFMT system which uses an Electron-Multiplying charge coupled device (EMCCD). More precisely, we report on the effects of EM gain and deep cooling of the camera on signal-to-noise ratio (SNR) and hence the sensitivity. Experimental results showed that these parameters, when used in conjunction, increased SNR by at least 3 fold.