Tuesday, 14 November 2017

Improved mesoscopic imaging - proven in uncleared tumor spheroids

Scientists at the Foundation for Research and Technology – Hellas (FORTH) in Heraklion, Greece have developed a significantly improved methodology for the optical imaging of optically opaque 3D cell cultures with enhanced resolution.

The so-called Phase-Retrieved Tomography (PRT) utilises a Light Sheet / Selective Plane Illumination Microscope (SPIM), Optical Projection Tomography and algorithms to solve challenges around autocorrelation.  PRT’s ability to permit 3D image reconstruction was tested on unfixed, uncleared multicellular tumor spheroids of 200 ┬Ám diameter for early stage necrosis using a fluorescent viability dye as a representative and practically relevant proof of concept for the technology.

In the model system individual tumor spheroids were grown in hanging-drop cultures, and further incubated with the addition of the far-red viability dye DRAQ7 for 24 hours to mark the presence of membrane-compromised dead cells. (open acess)

After a wash procedure a single spheroid was then transferred into CyGEL Sustain, a thermo-reversible hydrogel mountant compatible with living cells and tissues, and this combination placed in a FEP tube, which was sealed and placed in a 37 °C waterbath for the duration of the imaging procedure.

Reference: 
Ancora, D., Di Battista, D., Giasafaki, G., Psycharakis, S. E., Liapis, E., Ripoll, J., & Zacharakis, G. (2017). Phase-Retrieved Tomography enables Mesoscopic imaging of Opaque Tumor Spheroids.