Studying Cell-Cell Interactions in Facilitating Neuronal Maturation

Start with a co-culture of astrocytes and differentiating neuronal stem cell aggregates in differentiation media.

Astrocytes secrete signaling molecules that enhance the maturation and differentiation of stem cells into neurons and astrocytes.

Next, add a fixative to preserve the cell structure, followed by washing.

Add a detergent solution to permeabilize the cell membranes, then wash.

Introduce a blocking reagent to block nonspecific binding sites.

Add primary antibodies that specifically bind to protein markers of mature neurons and astrocytes. Wash to remove unbound antibodies.

Then, add green and red fluorescently labeled secondary antibodies and incubate. These antibodies bind to respective primary antibodies, providing distinct fluorescence to cells.

Introduce a nuclear dye to stain the nuclei blue.

Finally, observe the cells using a confocal microscope.

Within the aggregates, green fluorescent cells indicate mature neurons, while red fluorescent cells represent astrocytes. This confirms cell-cell interactions in facilitating neuronal maturation.