Three – Dimensional Culture For Continuous Long Term Live Cell Imaging

Conventional cell culture systems are primarily two dimensional (2D). They are low cost and robust, and ensure homogeneous cell proliferation due to equal access to nutrients. However, they do not accurately represent in vivo conditions, with only horizontal spreading, in case of adherent cells. Three dimensional (3D) culture systems are now becoming increasingly popular since the conditions in these systems are similar to those in vivo, due to the presence of a matrix. 3D systems also make it easier to image and track individual cells over long periods of time, especially suspension cells. Continuous long term live cell imaging using a 3D culture system can aid in the detection of minute morphological changes in cells which can serve as indicators of complex cellular processes. In our study, we assessed the suitability of a collagen-based 3D culture system for continuous long term live cell imaging with respect to the collagen concentrations, cell seeding numbers for adherent and suspension cells and the longest duration for imaging. A primary test for diffusion of dye through the gel was also conducted. We found collagen concentrations of 1.5 and 2.0 mg/ml seeded with cell numbers from 1× 105 and 3 × 105 cells/ml (depending on the cell type) to be optimal for imaging up to 120 hours. The primary dye diffusion test showed successful staining of cells through the matrix. Thus, we demonstrate the practicability of 3D culture for continuous long term live cell imaging of adherent and suspension cells.