We have screened Xenopus animal cap ectodermal cells during neural induction for protein kinases (PK) and protein tyrosine kinases (PTKs) after their renaturation in gels containing the polydispersed substrate poly(Glu, Tyr). Following guanidine hydrochloride denaturation, renaturation, and phosphorylation with [gamma-32P]ATP, kinase activity was detected by autoradiography. Incubation of gels in hot alkali after glutaraldehyde crosslinking completely eliminated the activity of non-PTK enzymes. This method is very sensitive and can be applied to development biology for the detection of PTKs in very small samples such as ectoderm explants dissected from animal caps in amphibian embryos. A large number of PTKs were visualized in uninduced and induced ectodermal cells. This procedure should be useful for investigating the role of PTKs in intracellular signaling during neural induction.
Animals Autoradiography