In Vivo Staining by Dis and Trisazo Dyes, Especially of Bone and Elastic Tissue

The localization and retention of dis and trisazo dyes in connective tissues and bone was studied in rats and rabbits. Chlorazol fast pink, 5% in 0.9% NaCl. was injected intraperitoneally, 25 mg/kg each day for 2 days in newborn, growing, mature and 17-18 day pregnant rats, and up to 5 days in young rabbits. The dye was also injected at different time intervals during the development of strontium-induced rickets in growing rats, and in animals with abscess walls following subcutaneous injection of 0.1 ml turpentine. Animals were killed at-intervals thereafter, and comparison of in vivo staining of 5% solutions of chlorazol fast pink, chlorantine fast red, chlorazol black E, chlorazol sky blue, chlorazol sky pink, chlorazol green, chlorazol violet, pontamine green and pontamine sky blue was made by intraperitoneal injection in rats. Soft and hard tissue specimens were embedded in polyester resin or in paraffin wax and sectioned at 5-7 μ. Chlorazol fast pink stained some connective tissues and growing bones. The main intensity of staining occurred within 24 hr and gradually decreased but was still detectable after 6 mo in elastic tissues. In thin plastic sections, colouration was brilliant, not in osteoid tissue, but at calcifying bone margins and in elastic fibres. Dye localized at calcifying bone margins was incorporated within calcified tissues and then subsequently lost through remodelling. Such staining was not seen in paraffin-embedded material. Dye uptake was greatly reduced in rachitic rats, and wide osteoid seams were coloured faint pink, but where calcification was still occurring, colouration was brilliant. Similarly collagenous tissue in abscess walls was only lightly stained, in contrast to brilliant colouration of elastic tissues and macrophages. Of the 9 dyes tested, only chlorazol fast pink and chlorazol sky blue stained bone and elastic tissue in vivo. This prolonged retention and staining by these 2 dyes, unlike the others, was associated with their presence in the proximal convoluted tubules of the kidney.