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Fig. 2. Tissue distribution of T protein in limb bud stage chick and mouse embryos. (A) Western analysis of T protein in dissected tissues using affinity-purified polyclonal antibody. T protein (47 kDa band) was detected in chick wing (W) and leg (L) buds and in somites (So) at stages (st) 18 and 20, as well as in wild-type (wt) E10 mouse forelimb (FL) and hindlimb (HL) buds. No T protein band was detected in head or trunk (including forelimb buds) from T-/- mouse embryos (negative control). Chick or mouse tailbud protein was loaded as a positive control. A ubiquitous 100 kDa protein was recognized by affinity-purified T antibody in every tissue evaluated. Protein loading was assessed by reprobing blots with {alpha}-tubulin antibody (bottom). (B) Immunohistochemical detection of T protein in transverse hindlimb section of E10.5 mouse embryo, showing expression in notochord (no) somitic dermomyotome (so) and limb mesoderm subjacent to AER. Affinity purified antibody was exhaustively pre-adsorbed against 100 kDa proteins prior to use. d, dorsal; v, ventral.