Fig 1.
Schematic of agarose microwell fabrication and cell seeding for preparation of glioma spheroids.
Fig 2.
Phase contrast images of representative U251 glioma spheroids prepared in microwells of 70, 150, 450, and 700 μm diameter.
Scale bars represent 200 μm.
Fig 3.
Characterization of U251 cells in spheroid and monolayer cultures.
A) Phase contrast and fluorescence images of U251 cells in the form of a spheroid produced in a 150 μm well (top row) and monolayer (bottom row). Green fluorescence represents the live cells and red fluorescence represents dead cells. Yellow dashed circles mark the border of a single hydrogel microwell. Scale bars represent 80 μm. B) Sub-G1 population of U251 glioma cells in spheroids (top) and monolayers (bottom).
Fig 4.
Effect of initial cell seeding density and hydrogel microwell size on the final size of U251 glioma spheroids.
Graphs show the volume of U251 spheroids, measured after 2 days in culture as a function of cell seeding density per well of the culture plate that contained agarose microwells with diameters of 70 μm (A), 150 μm (B), and 450 μm (C). Data points represent the mean of 10 independent experiments and error bars represent standard deviation (SD). Solid lines show the linear fit to the data.
Fig 5.
Growth rate of U251 spheroids in agarose microwells.
Graph shows the measured volume of U251 spheroids as a function of time in 450 μm-sized microwells (cell seeding densities of 6.0 × 105 cells/well). Data points represent the mean of 6 independent experiments and error bars show SD. Solid line shows the linear fit to data (y = 16.2x + 191.7, R2 = 0.9610).
Fig 6.
Effect of an anti-tumor agent Dp44mT on U251 spheroids.
A) Bar graph shows the volume of glioma spheroids upon exposure to 0 nM Dp44mT (control) and 100 nM Dp44mT for five days. Data points represent the mean of 10 independent experiments and error bars represent SD. * represents P < 0.01 vs. control (0 nM). The inset shows phase contrast images of the control and Dp44mT-treated spheroids. Sale bar represents 50 μm. B) Flow cytometry data demonstrate the sub-G1 population (i.e. apoptotic cell population) in control and Dp44mT-treated (100 nM) spheroids.