The binding of Evans blue to collagen and elastin in elastic tissue

The binding of Evans blue to collagen and elastin in rabbit aortic tissue and in bovine ligamentum nuchae was studied following circulation in vivo of the dye and incubation in vitro in Evans blue containing plasma, respectively. Using collagenase and elastase, the dye was liberated from both tissues corresponding to their different contents of collagen and elastin. Disc electrophoretic analysis of the liberated dye showed, that it migrated as free Evans blue indicating that the binding of the dye to the macromolecules was due to spatial interactions rather than to fixation at specific prosthetic groups. The capability of collagen and elastin to bind Evans blue was demonstrated with the isolated proteins; it was shown elastin had a higher affinity to the dye than collagen. Treatment of the blued tissue with hyaluronidase and Triton X-100 showed that binding to complex carbohydrates and dye accumulation in the aqueous intra- or extracellular space seems to be negligible.