Explant coculture

After incubation, fix and develop the explant coculture for lacZ expression in reporter cells as outlined in steps 14-19. 

Gently remove medium from the explant cocultures taking care not to dislodge the tissue explants. Gently wash the cocultures twice with PBS Fix the tissue and cells by incubating in fixative solution for 15 mm at room temperature... 

Alfalfa leaf explants from a highly regenerable clone of Regen-SY (19) were cocultured with A. tumefaciens containing each binary vector, and... 

Figure 2. Schematic view of collagen gel coculture of a trigeminal ganglion with a maxillary arch explant. Side view [A] and top view [B] of the culture dish.

Hgure 6. Chemorepulsion of mitral cell axons by septum. (A) Axon growth pattern of mitral cells in the olfactory bulb. Mitral cell axons grow laterally away from the septum and form the lateral olfactory tract in the lateral telencephalon. (B) Collagen gel coculture of olfactory bulb explants with septal explants. 

The F9-lacZ reporter assay provides a qualitative indication of retinoid release from tissue explants maintained m short term coculture with reporter cells. There are two important requirements that must be kept in mind in order to successfully use the assay, particularly when tissue samples are taken from developing embryos. The first requirement is that the tissue being studied must be viable under the culture conditions required for maintaining the reporter cell monolayer (see Note 7). The second requirement is that the development of the embryos must be timed in accordance with the plating of the reporter cells to ensure that tissue from the desired developmental stage is plated onto near-confluent reporter cell monolayers. 

Place a thin layer of a 50% glycerol solution in PBS over the cocultures and cover slip, taking care not to dislodge the tissue explants. Storage at 4°C is recommended and often intensifies the blue reaction product... 

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