Alizarin (madder) has been used in textile dyeing since early antiquity. In histology calcium-alizarin or calcium-alizarin red S compounds are often referred to as "lake" or "complex." Chemical and infrared spectroscopic data showed that these compounds are salts, not chelates. In dye chemistry the term lake denotes a poorly soluble or insoluble salt of a water-soluble dye. Salt formation between calcium deposits in tissues and alizarin or alizarin red S is indicated by the sensitivity of these compounds toward dilute acetic acid. Acid dyes for lakes, which do not contain chelating groups, also stained calcium deposits selectively. Alizarin stained calcium deposits intensely only around pH 12. Alizarin red S colored calcium deposits selectively around pH 9; neutral and acid dye solutions produced severe diffusion artifacts. Chemical data indicate that alizarin red S can react with calcium via its sulfonic acid and/or its OH groups.