Clearing and Staining Whole Fish Specimens for Simultaneous Demonstration of Bone, Cartilage, and Nerves

Abstract

Comparative anatomical studies of nerves alone, or combined with those of bone and cartilage, are rare in ichthyology because of lack of a reliable, repeatable protocol for preparing specimens that demonstrate all three character systems. We present a method for enzyme clearing and staining whole fish specimens for simultaneous demonstration of bone, cartilage, and nerves. Nerves are stained dark blue with Sudan Black B following alcian blue-alizarin red S staining. Our protocol for Sudan Black B staining differs from others in concentration of Sudan Black B solution, length of time in that solution, and method of differentiation.