Real architecture For 3D Tissue (RAFT™) culture system improves viability and maintains insulin and glucagon production of mouse pancreatic islet cells.

Szebeni GJ, Tancos Z, Feher LZ, Alfoldi R, Kobolak J, Dinnyes A, Puskas LG
Source: Cytotechnology
Publication Date: (2017)
Issue: 69(2): 359-369
Research Area:
Basic Research
Drug Discovery

After 3 days of maintenance, islets were diluted to plate an average 3 islets per well for subsequent analysis in 96-multiwell RAFTTM (3D) plates according to the instructions of the manufacturer (TAP Biosystems, Lonza, Cologne, Germany). Briefly, islets were dispensed in the chilled mixed collagen solution of the RAFTTM kit (containing 2.8 ml 10X MEM medium, 22.4 ml 2 mg/ml rat tail collagen type I, 1.624 ml neutralizing solution and 1.2 ml islets for 96-well tissue culture plate) according to the RAFT TM protocol. We plated 240 µl mixed collagen solution per well and incubated the plate at 37 °C to form a hydrogel for 15 min. Then we placed the RAFTTM absorbers on the top of the hydrogel in a laminar flow hood at room temperature (RT) for 15 min. After removing the absorbers we immediately loaded 200 µl MES medium per well. Medium was replaced on every third day.


There is an unmet medical need for the improvement of pancreatic islet maintenance in culture. Due to restricted donor availability it is essential to ameliorate islet viability and graft engraftment. The aim of this study was to compare the standard tissue culture techniques with the advanced Real Architecture For 3D Tissue (RAFT™) culture system in terms of viability and hormone production. Here, we first report that islets embedded in RAFT™ collagen type I advanced tissue culture system maintain their tissue integrity better than in monolayer and suspension cultures. The Calcein violet assay and Annexin V/propidium-iodide staining show higher cell viability in the RAFT™ culture system. Quantitative real-time PCR data showed that RAFT™ increases insulin expression after 18 days in culture compared to traditional methods. Enhanced insulin and glucagon production was further verified by immunofluorescent staining in a time-course manner. These results indicate that RAFT™ tissue culture platform can be a promising tool to maintain pancreatic islet spheroid integrity and culture islets for downstream high throughput pharmacological studies ex vivo.