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Objective: In biomanufacturing there is a need for quantitative methods to map cell viability and density inside 3D bioreactors to assess health and proliferation over time. Recently, noninvasive MRI readouts of cell density have been achieved. However, the ratio of live to dead cells was not varied. Herein we present an approach for measuring the viability of cells embedded in a hydrogel independently from cell density to map cell number and health., Methods: Independent quantification of cell viability and density was achieved by calibrating the 1 H magnetization transfer- (MT) and diffusion-weighted NMR signals to samples of known cell density and viability using a multivariate approach. Maps of cell viability and density were generated by weighting NMR images by these parameters post-calibration., Results: Using this method, the limits of detection (LODs) of total cell density and viable cell density were found to be 3.88 ×10 8 cells · mL -1 · Hz -1/2 and 2.36 ×10 9 viable cells · mL -1 · Hz -1/2 respectively., Conclusion: This mapping technique provides a noninvasive means of visualizing cell viability and number density within optically opaque bioreactors., Significance: We anticipate that such nondestructive readouts will provide valuable feedback for monitoring and controlling cell populations in bioreactors.

Objective: For tissue engineering, there is a need for quantitative methods to map cell density inside three-dimensional (3-D) bioreactors to assess tissue growth over time. The current cell mapping methods in 2-D cultures are based on optical microscopy. However, optical methods fail in 3-D due to increased opacity of the tissue. We present an approach for measuring the density of cells embedded in a hydrogel to generate quantitative maps of cell density in a living, 3-D tissue culture sample., Methods: Quantification of cell density was obtained by calibrating the 1 H T 2 , magnetization transfer (MT) and diffusion-weighted nuclear magnetic resonance (NMR) signals to samples of known cell density. Maps of cell density were generated by weighting NMR images by these parameters post-calibration., Results: The highest sensitivity weighting arose from MT experiments, which yielded a limit of detection (LOD) of [Formula: see text] cells/mL/ √{Hz} in a 400 MHz (9.4 T) magnet., Conclusion: This mapping technique provides a noninvasive means of visualizing cell growth within optically opaque bioreactors., Significance: We anticipate that such readouts of tissue culture growth will provide valuable feedback for controlled cell growth in bioreactors.