This video demonstrates how to encapsulate and culture cancer cells in PuraMatrix, a commercially available self assembling peptide gel.
Technical Notes:
Materials:
BD PuraMatrix RAD16 Peptide Hydrogel-1% (Cat # 354250): The total peptide concentration of the undiluted mixture is 10 mg/ml.
All steps must be performed under aseptic conditions.
Procedure:
Representative Results:
Figure 1. OVCAR-5 cells were encapsulated as described in the above protocol. Images were acquired using an inverted Olympus FV1000 microscope equipped with Spectra-Physics DeepSee Ti:Sapphire laser tuned to 750 nm. Imaging sessions were conducted on days 1, 3, 5 and 7 post-plating. A long working distance, water dipping 40x objective (Olympus LUMPLFLN 0.8 NA) was used to image through the PuraMatrix. Three-dimensional volumes were collected by acquiring image stacks in 1.47 μm axial steps. Two non-descanned detectors collected the autofluorescence emission using violet (440-490 nm) and green (510-550 nm) bandpass filters. The final images and depth stack movies were processed using ImageJ by combining both detection channels. Please click here to see a larger version of figure 1.