Assessment of chitinase kinetics and mechanism in vitro has been hampered by lack of suitable substrates. We have previously reported rapid linear initial chitinase velocity with chitin substrate isolated from insect larval cuticle. Such chitin is shown to be fibrous in the light microscope. Methods are described for preparing fibrous chitins from any animal source including calcified carapaces. Evidence is given that chitin native fine structure in situ is maintained by structural proteins which in the fibrous chitin isolates are functionally replaced by covalently bound ester groups. Chitin fiber analogues thus reconstructed appear to have retained their native fine structure.