This video demonstrates a technique for immunostaining cells encapsulated in a 3D hydrogel. The hydrogel culture containing neural progenitor cells is fixed, permeabilized, and treated with a blocking solution to prevent non-specific immunostaining. A primary antibody cocktail is added to bind specific proteins, followed by fluorescently labeled secondary antibodies and a nuclear stain. The sample is then mounted on a slide and observed under a confocal microscope.
Protocol
1. Cell Encapsulation in 3D Elastin-like Protein (ELP) Hydrogels Preparation of silicone molds Use a biopsy punch with the desired diameter to create holes in a 0.5 mm thick silicone sheet and cut out a square around each hole. Repeat for the number of molds desired. NOTE: The diameter and thickness of the molds can be adjusted for the particular application and cell culture conditions. In practice, for cell cultures of 50 x 10…