A safer and more sensitive substitute for diamino-benzidine in the light microscopic demonstration of retrograde and anterograde axonal transport of HRP.

The commonly used benzidine derivatives such as diaminobenzidine, o-dianisidine and benzidine dihydrochloride are apparently carcinogenic. Available information, however, indicates that tetramethyl benzidine (TMB) is safer in this regard, and we have therefore developed a procedure in which TMB is used as a substrate for the detection of intra-axonal transport of HRP. The TMB procedure is simple, reliable and does not produce crystals if properly executed. The staining procedure, which takes about 15 min, produces a blue reaction product which is easily detected in bright-field microscopy.