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Three-dimensional, label-free cell viability measurements in tissue engineering scaffolds using optical coherence tomography



Greta Babakhanova, Anant Agrawal, Deepika Arora, Allison Horenberg, Jagat Budhathoki, Joy Dunkers, Joe Chalfoun, Peter Bajcsy, Carl Simon Jr.


In the field of tissue engineering, 3D scaffolds and cells are often combined to yield constructs that are used as therapeutics to repair or restore tissue function in patients. Viable cells are required to achieve the intended mechanism of action for the therapeutic, where the live cells may build new tissue or may release factors that induce tissue regeneration. Thus, there is a need to be able to reliably measure cell viability in 3D scaffolds as a quality attribute of a tissue-engineered medical product. Here, we utilized a noninvasive, label-free, 3D optical coherence tomography (OCT) method to rapidly (2.5 min) image large sample volumes (1 mm3) to assess cell viability and distribution within scaffolds. A model scaffold-cell system consisting of a polysaccharide-based hydrogel seeded with human Jurkat cells was used to assess OCT imaging. Four test systems were used: hydrogel seeded with live cells, hydrogel seeded with heat-shocked or fixed dead cells and hydrogel without any cells. Time series OCT images demonstrated changes in the time-dependent speckle patterns due to refractive index (RI) variations within live cells that were not observed for pure hydrogel samples or hydrogels with dead cells. The changes in speckle patterns were used to generate live-cell contrast by image subtraction. In this way objects with large changes in RI could be binned as live cells and objects with smaller changes in RI could be binned as non-live cells, which would include dead cells, debris or hydrogel inclusions (bubble or inhomogeneities). Using this approach, on average, OCT imaging measurements counted 326±52 live cells per 0.288 mm3 for hydrogels that were seeded with 288 live cells in 0.288 mm3. Considering the substantial uncertainties in fabricating the scaffold-cell constructs, such as the error from pipetting and counting cells, a 13% difference in the live-cell count is within a reasonable error. Additionally, the 3D distribution of live cells was mapped within a hydrogel scaffold to assess the uniformity of their distribution across the volume. Our results demonstrate a real-time, noninvasive method to rapidly assess cell viability and spatial distribution throughout a 3D scaffold that could be useful for assessing tissue-engineered medical products.
Journal of Biomedical Materials Research Part A


cell viability, tissue engineering, 3D scaffolds, noninvasive imaging, optical coherence tomography


Babakhanova, G. , Agrawal, A. , Arora, D. , Horenberg, A. , Budhathoki, J. , Dunkers, J. , Chalfoun, J. , Bajcsy, P. and Simon Jr., C. (2023), Three-dimensional, label-free cell viability measurements in tissue engineering scaffolds using optical coherence tomography, Journal of Biomedical Materials Research Part A, [online],, (Accessed April 21, 2024)
Created March 14, 2023, Updated March 19, 2023