It has been proposed that gradients of environmental cues direct the proximal growth of pioneer axons in embryonic insect legs. Hybridoma techniques have been used to produce 3 monoclonal antibodies (mAbs) that bind to components associated with the basal lamina/extracellular matrix that are non-uniformly distributed along the proximal-distal axis of cockroach legs at the time of pioneer axon growth. Two of these mAbs, PROD-1 and PROD-2, label the proximal parts of the leg more intensely than the distal ends. The other mAb, DIP-1, has the reverse pattern of binding with the distal parts of the leg labeled more intensely. The graded distribution of these antigens only occurs just prior to and during the growth period of the Ti1 pioneer axons. Western blot analyses and immunoprecipitations have identified the protein antigens recognized by these mAbs. The spatial and temporal distributions of these molecules in the legs and the CNS make them good candidates for environmental guidance cues of pioneer axon growth.