Calcium is present in large quantities in the human body, constituting the skeleton as a scaffolding to support the body, and also plays an important role in secretion, transportation, muscle contraction, and nerve conduction. Calcium exists in two forms in the body, one is ionized calcium, which exists within the blood circulation and is known as blood calcium, and the other is bound calcium, which is bound to proteins, carbonic acid, or phosphoric acid and deposited in the tissues. Calcium normally permeates all tissues and cells except bones and teeth, and does not normally appear in a solid state in the tissues, but in some cases it precipitates as a solid and settles in the tissues, which is pathologically calcium-salt deposits, the deposited calcium salts being mainly calcium phosphate and to a lesser extent calcium carbonate. Calcium salts are usually monorefractive, but calcium oxalate is birefractive. When stained with HE, calcium generally appears as a purplish-blue color, and a number of dyes can form chelates with calcium, including alizarin red S, red violet, and nuclear solid red.

Alizarin Red S is an anthraquinone derivative, a sodium salt of alizarin sulfonate, which chelates with calcium salts in calcium carbonate or calcium phosphate to form an orange-red complex. Alizarin Red S tends to stain small amounts of sediment to give more reliable results, and is often used in combination with solid green or Mayer's Hematoxylin stains to form an orange-red precipitate, which is suitable for staining small amounts of calcium salts in tissue.